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Information on EC 1.14.15.1 - camphor 5-monooxygenase and Organism(s) Pseudomonas putida and UniProt Accession P00183
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1.14.15.1 camphor 5-monooxygenaseIUBMB Comments
A heme-thiolate protein (P-450). Also acts on (-)-camphor and 1,2-campholide, forming 5-exo-hydroxy-1,2-campholide.
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Word Map
- 1.14.15.1
- heme
-
ferrous
-
substrate-free
-
camphor-bound
-
substrate-bound
-
low-spin
-
high-spin
- dioxygen
-
p450terp
-
soret
- chloroperoxidase
- p450eryf
-
self-sufficient
- adrenodoxin
- aromaticivorans
-
monoxygenase
- novosphingobium
-
monooxygenation
-
ferryl
-
thiolate-ligated
-
i-helix
-
oxyferrous
- analysis
- synthesis
-
electron-electron
- biotechnology
The taxonomic range for the selected organisms is: Pseudomonas putida
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota
Reaction Schemes
+
reduced putidaredoxin
+
=
+
+
Synonyms
p450cam, cytochrome p450cam, cyp101, cytochrome p450(cam), cyp101d1, cyp101a1, camphor hydroxylase, cyp101d2, cyt p450cam, cytochrome p-450-cam, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
cytochrome p450cam
-
P450cam
-
2-bornanone 5-exo-hydroxylase
-
-
-
-
bornanone 5-exo-hydroxylase
-
-
-
-
camphor 5-exo-hydroxylase
-
-
-
-
camphor 5-exo-methylene hydroxylase
-
-
-
-
camphor 5-exohydroxylase
-
-
-
-
Camphor 5-monooxygenase
-
-
-
-
camphor hydroxylase
-
-
-
-
camphor methylene hydroxylase
-
-
-
-
cytochrome p450cam
d-camphor monooxygenase
-
-
-
-
D-camphor-exo-hydroxylase
-
-
-
-
haem mono-oxygenase CYP101
-
-
-
-
methylene hydroxylase
-
-
-
-
methylene monooxygenase
-
-
-
-
oxygenase, camphor 5-mono-
-
-
-
-
P450cam
cytochrome p450cam
-
-
-
-
cytochrome p450cam
-
-
P450cam
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
binding of putidaredoxin forces selection of the active conformation of enzyme
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
active binding of substrate camphor, analysis by density functional theory calculations, residue Tyr96 is important forming a strong hydrogen bond, catalytic cycle of cytochrome P450, the strong hydrogen bonding is not affected by the enzyme's environment, reaction mechanism, overview
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
rebound mechanism of C-H hydroxylation by P450, molecular dynamics
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
the active site structure changes due to putidaredoxin-enzyme interaction involving salt bridge and hydrogen bonding network between Arg112, His355, Leu356, and the heme ligand Cys357
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
proposed catalytic cycle for cytochrome P450, a ferryl-oxo Pi-cation porphyrin radical, is the putative oxidant that reacts directly with substrate, overview. The axial ligand to the heme iron, a cysteine thiolate, is generally believed to control P450 reactivity
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
proposed catalytic cycle for cytochrome P450, overview
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
reaction mechanism
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
multi-component mixed function oxidase, consisting of putidaredoxin reductase, putidaredoxin and cytochrome P450cam, heme-thiolate protein
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
model for putidaredoxin activity, primary role of putidaredoxin is to prevent uncoupling by enforcing conformations of enzyme that prevent loss of substrate and to enforce conformations that permit efficient proton transfer
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
oxygen-transfer reaction via a monooxygenation mechanism
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
putidaredoxin acts as a shuttle for transport of electrons from putidaredoxin reductase to enzyme
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
the acvtive conformation of enzyme is stabilized by binding of the substrate at the active site
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
K+ plays an important role in substrate binding and structural and conformational stability of the enzyme
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
mechanism of O2 activation: binding of O2 to ferrous P450cam to yield the ferric-superoxo form, oxyP450cam, followed by an irreversible, long-range electron transfer from putidaredoxin to reduce the oxyP450cam, camphor is bound to all enzyme forms, overview
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
reaction mechanism and cycle of the enzyme including postulated intermediates, detailed overview
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
reaction mechanism involving a heme ring and residue Asp297, modelling of the hydroxylation of camphor and the hydrogen abstraction from heme using combined quantum mechanical/molecular mechanical method
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
reaction mechanism, and second reductive step of the mechanism of interaction and electron transfer, overview
-
(+)-camphor + reduced putidaredoxin + O2 = (+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
via H2O2 and a nucleophilic intermediate, the substrate can modulate the properties of both the monoxygenase active-oxygen intermediates and the proton-delivery network that encompasses them, the catalytic cycle, overview
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
redox reaction
-
-
-
-
oxidation
-
-
-
-
reduction
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
(+)-camphor,reduced putidaredoxin:oxygen oxidoreductase (5-hydroxylating)
A heme-thiolate protein (P-450). Also acts on (-)-camphor and 1,2-campholide, forming 5-exo-hydroxy-1,2-campholide.
CAS REGISTRY NUMBER
COMMENTARY
9030-82-4
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
(+)-alpha-pinene + putidaredoxin + O2
(+)-cis-verbenol + (+)-myrtenol + (+)-verbenone + oxidized putidaredoxin + H2O
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-camphor + O2 + reduced putidaredoxin
exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-camphor + reduced putidaredoxin + NADH + H+ + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + NAD+ + H2O
(+)-camphor + reduced putidaredoxin + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(+)-camphor + reduced putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(1R)-(+)-camphor + reduced putidaredoxin + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(1R)-5,5-difluorocamphor + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
(1R)-5-exo-methoxycamphor + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
(1R)-5-methylenylcamphor + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
(1R)-camphor + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
(1R)-norcamphor + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
(1S)-camphor + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
1-ethyl-2-methylbenzene + reduced putidaredoxin + O2
? + oxidized putidaredoxin + H2O
the S-enantiomer is preferred by the wild-type enzyme
-
-
?
1-ethyl-3-methylbenzene + reduced putidaredoxin + O2
? + oxidized putidaredoxin + H2O
the S-enantiomer is preferred by the wild-type enzyme
-
-
?
1-ethyl-4-methylbenzene + reduced putidaredoxin + O2
? + oxidized putidaredoxin + H2O
the S-enantiomer is preferred by the wild-type enzyme
-
-
?
3-chloroindole + O2 + reduced putidaredoxin
isatin + H2O + Cl- + oxidized putidaredoxin + ?
no substrate of wild-type, substrate of mutants E156G/V247F/V253G/F256S, T56A/N116H/D297N and G60S/Y75H
-
-
?
3-chloroperbenzoic acid + O2 + reduced putidaredoxin
?
compound I, ferryl iron plus a porphyrin pi-cation radical (Fe(IV)=O/Por(+)), and compound ES, Fe(IV)=O/Tyr(), in reactions of substrate-free ferric enzyme with 3-chloroperbenzoic acid, compound ES arises by intramolecular electron transfer from nearby tyrosines to the porphyrin pi-cation radical of compound I, active site changes influence electron transfer from nearby tyrosines and affect formation of intermediates, the tyrosyl radical is assigned to Tyr96 for wild type or to Tyr75 for the Y96F variant, overview
-
-
?
adamantanone + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
camphane + reduced putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
DL-camphor + reduced putidaredoxin + NADH + H+ + O2
exo-5-hydroxycamphor + oxidized putidaredoxin + NAD+ + H2O
-
-
-
?
isoborneol + reduced putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
norcamphor + reduced putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
thiocamphor + reduced putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
?
(+)-alpha-pinene + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-camphor + O2 + reduced putidaredoxin
exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-camphor + reduced putidaredoxin + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-exo-5-hydroxycamphor + reduced putidaredoxin + O2
5-oxocamphor + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-camphor + putidaredoxin + O2
5-exo-(1R)-hydroxycamphor + oxidized putidaredoxin + H2O
(1R)-camphor + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-camphor enol ether + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-camphor N-methyl imine + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-camphor oxime + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-endo-borneol allyl ether + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-endo-borneol methyl ether + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-endo-borneol propyl ether + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(1R)-iso-borneol methyl ether + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
(4S)-limonene + putidaredoxin + O2
?
-
the 7-position is the major site of hydroxylation by P450cam
-
-
?
(R)-2-ethylhexanol + reduced putidaredoxin + O2
(R)-2-ethylhexanoic acid + oxidized putidaredoxin + H2O
-
-
-
-
r
(R)-3-ethylhexanol + putidaredoxin + O2
2-ethylhexanoic acid + 2-ethyl-1,2-hexanediol + 2-ethyl-1,3-hexanediol + 2-ethyl-1,4-hexanediol + oxidized putidaredoxin + H2O
-
-
ratio: 50:13:15:8
?
(R)-exo-5-hydroxycamphor + O2 + reduced putidaredoxin
2,5-diketocamphane + oxidized putidaredoxin + H2O
-
-
-
-
?
(S)-2-ethylhexanol + reduced putidaredoxin + O2
(S)-2-ethylhexanoic acid + oxidized putidaredoxin + H2O
-
-
-
-
r
(S)-3-ethylhexanol + putidaredoxin + O2
2-ethylhexanoic aicd + 2-ethyl-1,2-hexanediol + 2-ethyl-1,3-hexanediol + 2-ethyl-1,4-hexanediol + oxidized putidaredoxin + H2O
-
the (S)-isomer is turned over 1.4times faster than the (R)-isomer
ratio: 15:53:28:10
?
1,2,4,5-tetrachlorobenzene + putidaredoxin + O2
2,3,5,6-tetrachlorophenol + oxidized putidaredoxin + H2O
-
-
-
?
1,2-campholide + putidaredoxin + O2
5-exo-hydroxy-1,2-campholide + oxidized putidaredoxin + H2O
1,2-dibromo-3-chloropropane + O2 + reduced putidaredoxin
1-bromo-3-chloroacetone + allyl chloride + H2O + putidaredoxin + Br-
-
dehalogenation, bromochloroacetone is the major conversion product when the incubation medium is saturated with oxygen, while allyl chloride is the sole product in the absence of oxygen
a number of bromochloropropene are also formed to a minor extent by an elimination mechanism, product determination
-
?
1,2-dichlorobenzene + putidaredoxin + O2
2,3-dichlorophenol + 3,4-dichlorophenol + oxidized putidaredoxin + H2O
-
-
-
?
1,3,5-trichlorobenzene + putidaredoxin + O2
2,4,6-trichlorophenol + oxidized putidaredoxin + H2O
-
-
-
?
1,3,5-trichlorobenzene + putidaredoxin + O2
? + oxidized putidaredoxin + H2O
-
-
-
-
?
1,3-dichlorobenzene + putidaredoxin + O2
2,6-dichlorophenol + 2,4-dichlorophenol + 2,5-dichlorophenol + 2,3-dichlorophenol + oxidized putidaredoxin + H2O
-
-
-
?
1,4-dichlorobenzene + putidaredoxin + O2
2,5-dichlorophenol + oxidized putidaredoxin + H2O
-
-
-
?
1-dehydrocamphor + putidaredoxin + O2
exo-5,6-epoxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
3-chloroperbenzoic acid + O2 + reduced putidaredoxin
?
-
reaction mechanism of substrate-free ferric cytochrome P450cam, via FeIV-O plus porphyrin Pi-cation radical, overview
-
-
?
5,5-difluorocamphor + putidaredoxin + O2
5,5-difluoro-9-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
5-exo-bromocamphor + putidaredoxin + O2
5-ketocamphor + oxidized putidaredoxin + Br- + H2O
-
(+)- and (-)-enantiomer
-
?
benzo[a]pyrene + putidaredoxin + O2
3-hydroxybenzo[a]pyrene + oxidized putidaredoxin + H2O
-
-
-
-
?
ethylbenzene + putidaredoxin + O2
1-phenylethanol + oxidized putidaredoxin + H2O
-
at 5% of the reaction with (+)-camphor
ratio of (R)- to (S)-1-phenylethanol produced depends on mutant form
?
fluoranthene + putidaredoxin + O2
3-fluoranthol + oxidized putidaredoxin + H2O
-
-
-
-
?
indole + O2 + reduced putidaredoxin
3-hydroxyindole + oxidized putidaredoxin + H2O
-
no substrate of the wild-type enzyme, but a good substrate for Y96 mutants, mutant screening, overview
3-hydroxyindole undergoes spontaneous air oxidation to produce the insoluble dye indigo
-
?
phenanthrene + putidaredoxin + O2
1-phenanthrol + 2-phenanthrol + 3-phenanthrol + 4-phenanthrol + oxidized putidaredoxin + H2O
-
-
-
-
?
pyrene + putidaredoxin + O2
1-pyrenol + 2-pyrenol + 1,6-pyrenequinone + 1,8-pyrenequinone + oxidized putidaredoxin + H2O
-
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
analysis of protein-protein interactions between enzyme and cofactor, overview
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
binding of camphor is strongly dependent on the concentration of alcohols, alcohol expels camphor out of the heme cavity of the enzyme by affecting tertiary structure of Cyt P450cam as well as by modifying the solubility properties of camphor in aqueous medium, overview
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
regio- and stereoselective C-H bond hydroxylation, rebound mechanism, overview
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
though the active site of the enzyme resides deep inside the protein matrix, the substrate is recognized at the surface of the enzyme and directed towards the active site through the access channel. The threonine 192 that resides on the F-G loop and directed towards the putative substrate access channel of the enzyme, plays an important role in recognition of the substrate at the surface of the enzyme
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
the 7-propionate side chain plays a role in maintaining the high affinity of cytochrome P450cam for its substrate, the Asp297 and Gln322 residues are capable of undergoing a 7-propionate-associated conformational change in the protein interior
-
-
?
(+)-camphor + reduced putidaredoxin + NADH + H+ + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + NAD+ + H2O
-
-
-
?
(+)-camphor + reduced putidaredoxin + NADH + H+ + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + NAD+ + H2O
the catalytic cycle of P450cam requires two electrons, both of which are donated by putidaredoxin (Pdx), a ferredoxin containing a [2Fe-2S] cluster, structures of the Pdx-P450cam complex, overview
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
671535, 672100, 672115, 672742, 672760, 673037, 673071, 674152, 674438, 674474, 674516, 674959, 675234, 676280
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
terminal monooxygenase in a three-component camphor-hydroxylating system from Pseudomonas putida, the reaction cycle requires two distinct electron transfer processes from the [2Fe-2S] containing putidaredoxin to P450cam
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
determination of appearance of transient intermediates at 3°C by double mixing rapid scanning stopped-flow spectroscopy, electron transfer from reduced putidaredoxin gives high spin deoxyferrous P-450, substrate-free enzyme also binds the cofactor, but the cofactor does not deliver the electrons to the substrate-free oxyferrous enzyme, binding structure of camphor-bound oxyferrous P450-CAM with reduced putidaredoxin, functional implications of the formation of the perturbed oxyferrous intermediate, overview
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
mechanism of camphor hydroxylation incorporating an NADH-regeneration system, overview
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
regio- and stereospecific hydroxylation
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
second reductive step of the mechanism of interaction and electron transfer, overview
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
wild-type dioxygen complex structure: high occupancy and a ordered structure of the iron-linked dioxygen and two 'catalytic' water molecules that form part of a proton relay system to the iron-linked dioxygen, Thr252 accepts a hydrogen bond from the hydroperoxy (Fe(III)-OOH) intermediate that promotes the second protonation on the distal oxygen atom, leading to O-O bond cleavage and compound I formation
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
wild-type enzyme, but not mutant T252A
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
the reduced enzyme exhibits lower-amplitude motions of secondary structural features than the oxidized enzyme on all of the time scales accessible, and these differences are more pronounced in regions of the enzyme involved in substrate access to the active site (B' helix and beta3 and beta5 sheets) and binding of putidaredoxin (C and L helices), the iron-sulfur protein that acts as the effector and reductant of CYP101 in vivo
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
conformational change in CYP101 upon binding of putidaredoxin that re-orients bound camphor appropriately for hydroxylation
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
ferric hydroperoxo complex, elusive reactive species of cytochrome P450cam, and the hydroxo intermediate (formed during camphor hydroxylation) in the catalytic cycle of cytochrome P450cam all have a doublet ground state which have a pronounced multiconfigurational character in the case of compound I and the hydroxo intermediate
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
the Cpd II-like species is ineffective at hydroxylating camphor, but can be readily reduced by ascorbate to ferric P450cam, which can then bind camphor to form the high-spin heme
-
-
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
-
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
the (+)- and (-)-enantiomers serve as substrates
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
when deuterated at either 5-exo- or 5-endo-position, only 5-exo-hydroxycamphor is the product
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + reduced putidaredoxin + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
-
?
(+)-camphor + reduced putidaredoxin + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
-
r
(+)-camphor + reduced putidaredoxin + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
the enzyme shows regio- and stereospecific hydroxylation. Activation and cleavage of the oxygen molecule in the P450cam catalytic cycle is accompanied by two electron transfers from putidaredoxin. Ferric P450cam can accept the first electron from diverse chemical reductants and putidaredoxin homologues, but the second requires putidaredoxin as donor
-
-
?
(+)-camphor + reduced putidaredoxin + O2
borneol + oxidized putidaredoxin + H2O
-
under low oxygen conditions borneol is formed instead of 5-ketocamphor
-
-
?
(+)-camphor + reduced putidaredoxin + O2
borneol + oxidized putidaredoxin + H2O
-
the hydroxylation reaction proceeds via a catalytic cycle in which the reduction of dioxygen is coupled to the oxidation of the substrate. A key intermediate in the catalytic cycle is the iron-oxo species (Fe(IV)=O)
-
-
?
(+)-camphor + reduced putidaredoxin + O2
borneol + oxidized putidaredoxin + H2O
-
under low oxygen conditions
-
-
?
(1R)-camphor + putidaredoxin + O2
5-exo-(1R)-hydroxycamphor + oxidized putidaredoxin + H2O
-
putidaredoxin transfers electrons from NADH to P450cam in a coupled assay method
-
-
?
(1R)-camphor + putidaredoxin + O2
5-exo-(1R)-hydroxycamphor + oxidized putidaredoxin + H2O
-
putidaredoxin transfers electrons from NADH to P450cam
-
-
?
1,2-campholide + putidaredoxin + O2
5-exo-hydroxy-1,2-campholide + oxidized putidaredoxin + H2O
-
-
-
?
1,2-campholide + putidaredoxin + O2
5-exo-hydroxy-1,2-campholide + oxidized putidaredoxin + H2O
-
-
-
?
5-methylenyl-camphor + O2 + reduced putidaredoxin
?
-
wild-type enzyme and mutant T252A, in absence of the primary oxidant species of P450, the precursor species FeOOH can effect double bond activation of 5-methylenyl-camphor initiated by a homolytic cleavage of the O-O-bond and formation of an OH radical bound to the secondary oxidant by hydrogen bonding interaction, overview
-
-
?
peracetic acid + O2 + reduced putidaredoxin
?
-
reaction mechanism of substrate-free ferric cytochrome P450cam, via FeIV-O plus tyrosyl radical, overview
-
-
?
additional information
?
-
effects of heme environment on the hydrogen abstraction reaction of camphor in catalysis, overview
-
-
?
additional information
?
-
release of the substrate is caused both due to increased solubility of the substrate in solution in presence of alcohol and due to change in the tertiary structure of the active site of the enzyme, addition of alcohols to cytochrome P450cam causes a small change in the secondary structural elements but a significant change in the tertiary structural organization of the enzyme
-
-
?
additional information
?
-
an additional coupling pathway transpires during H2O2 shunting of the cycle of wild-type P450cam and in mutant T252A. The reaction starts with the FeIII(O2H2) intermediate, which transforms to Cpd I via a O-O homolysis/H-abstraction mechanism. The substrate 5-methylenylcamphor prevents H2O2 release, while the protein controls the FeIII(O2H2) conversion to Cpd I by nailing through hydrogen-bonding interactionsthe conformation of the HO radical produced during O-O homolysis. This conformation prevents HO readical attack on the porphyrins meso position, as in heme oxygenase, and prefers H-abstraction from FeIVOH thereby generating H2O + Cpd I. Cpd I then performs substrate oxidations
-
-
?
additional information
?
-
analysis of the molecular substrate recognition of wild-type and mutant P450cams by infrared (IR) spectroscopy, differing conformational heterogeneity in the active site of the P450cam variants and changes in heterogeneity upon binding of different substrates likely contribute to their variable affinities via a conformational selection mechanism, overview. Although P450cam is relatively specific for camphor, it also recognizes a number of small, camphor-like substrates, albeit with variable affinity. Substrate binding and active site structure, overview
-
-
?
additional information
?
-
catalytic turnover in P450cam requires the enzymes putidaredoxin (Pdx) and putidaredoxin reductase (Pdr), which mediate electron transfer from NADH to heme, the process is tightly coupled to substrate hydroxylation
-
-
?
additional information
?
-
comparison of substrate binding and catalytic ability of wild-type enzyme with putidareductase and putidaredoxin (ternary complex), and recombinant fusion enzyme P450cam-putidareductase with putidaredoxin, overview. The wild-type and mutant systems show comparable activity with camphor. Further oxidation of 5-exo-hydroxycamphor to 5-oxo-camphor by the fusion enzyme is 39% lower than for the native system
-
-
?
additional information
?
-
enzyme 450cam binds camphor and converts from the closed to open conformation upon binding putidaredoxin, the binding thermodynamics of Pdx differ when the conformation of P450cam is held in different states, thermodynamic analysis, overview
-
-
?
additional information
?
-
enzyme P450cam catalyzes the regioselective hydroxylation of d-camphor to 5-exo-hydroxycamphor. P450cam hydroxylates thiocamphor with a regioselectivity lower than camphor but higher than norcamphor. The high regioselectivity of camphor hydroxylation, the hydrogen bond (H-bond) between the camphor ketone and the side chain of active site residue Y96 influences the local electrostatics of the active site but has little effect on either the relative populations of conformational states or the nature of the energy landscapes within the conformations. Infrared spectrometric analysis of enzyme-substrate intercations, overview
-
-
?
additional information
?
-
enzyme P450cam produces benzyl alcohols with only moderate enantioselectivities, enantioselective benzylic hydroxylation catalysed by P450 monooxygenases, enantioselectivity of a library of active-site mutants of chimeric P450cam-RhFRed towards the benzylic hydroxylation of structurally related regioisomers of ethylmethylbenzene, molecular dynamics simulations and computational molecular modelling, overview
-
-
?
additional information
?
-
putidaredoxin (Pdx) and putidaredoxin reductase (PdR) are expressed in Escherichia coli strain BL21 from a pMG211 plasmid as C-terminally His6-tagged proteins, and purified by nickel affinity chromatography and gel filtration
-
-
?
additional information
?
-
-
modelling of the hydroxylation of camphor using crystal structure, PDB code 1DZ9, and combined quantum mechanical/molecular mechanical method, heme propionate side chains are not involved in catalysis, Asp297 is important for the reaction mechanism, overview
-
-
?
additional information
?
-
-
relative stability of dibromochloropropane and products, overview
-
-
?
additional information
?
-
-
the properties and reactivity of the oxyheme and of both the primary and the annealed intermediates are modulated by a bound substrate, including alterations in the properties of the heme center, the presence of any alternative substrate increases the lifetime of hydroperoxoferri-P450cam no less than about 20fold, especially 5-methylenyl-camphor
-
-
?
additional information
?
-
-
substrate specificities of wild-type and mutant enzymes, overview
-
-
?
additional information
?
-
-
under conditions of low oxygen, Pseudomonas putida cells and the isolated P450cam reduce camphor to borneol, product analysis by GC-MS
-
-
?
additional information
?
-
-
the P450FeIII-OOH intermediate must be the oxidizing species. The mechanism of hydrogen peroxide binding to the substrate-free form of P450cam is mainly governed by the ability of H2O2 to undergo deprotonation at the hydroxo ligand coordinated to the iron(III) center under conditions of pH > pKP450
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-camphor + reduced putidaredoxin + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
(+)-camphor + reduced putidaredoxin + O2
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
-
?
(+)-camphor + reduced putidaredoxin + O2
borneol + oxidized putidaredoxin + H2O
-
under low oxygen conditions borneol is formed instead of 5-ketocamphor
-
-
?
(1R)-camphor + putidaredoxin + O2
5-exo-(1R)-hydroxycamphor + oxidized putidaredoxin + H2O
-
putidaredoxin transfers electrons from NADH to P450cam in a coupled assay method
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
671535, 672100, 672115, 672742, 672760, 673037, 673071, 674152, 674438, 674474, 674516, 674959, 675234, 676280
-
-
?
(+)-camphor + O2 + reduced putidaredoxin
(+)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
terminal monooxygenase in a three-component camphor-hydroxylating system from Pseudomonas putida, the reaction cycle requires two distinct electron transfer processes from the [2Fe-2S] containing putidaredoxin to P450cam
-
-
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
(+)-camphor + putidaredoxin + O2
(R)-exo-5-hydroxycamphor + oxidized putidaredoxin + H2O
-
-
i.e. (+)-exo-5-hydroxycamphor
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
cytochrome b5
a cytochrome P450 enzyme, cytochrome b5 is bound in the reduced CYP101-camphor-carbon monoxide complex, cytochrome b5 perturbs many of the same resonances in the complex as Pdx, including those for residues involved in substrate access to and orientation within the active site of CYP101, chemical shifts, overview
-
cytochrome
-
state of cytochrome that is two equivalents of oxidation greater than the ferric form (Cpd I species), state of cytochrome that is one equivalent of oxidation greater than the ferric form (Cpd II species), two-electron-oxidized state of P450 or peroxidases containing both an oxoferryl center [FeIV=O] and either a tryptophanyl or tyrosyl radical, analogous to Cpd ES in cytochrome c peroxidase (Cpd ES species)
-
additional information
catalytic turnover in P450cam requires the enzymes putidaredoxin (Pdx) and putidaredoxin reductase (Pdr), which mediate electron transfer from NADH to heme, the process is tightly coupled to substrate hydroxylation
-
cytochrome P450
-
-
cytochrome P450
the putidaredoxin reductase-camphor monooxygenase fusion enzyme shows the 450 nm Soret band characteristic of the CYP superfamily
-
heme
structural, electronic, and catalytic properties of cytochrome P450cam are subtly altered when the cysteine that coordinates to the heme iron is replaced with a selenocysteine
putidaredoxin
-
671612, 672083, 673054, 674197, 675233, 676941, 701533, 701902, 702698, 704181, 744343, 744365, 744704, 745613, 746282
-
putidaredoxin
analysis of protein-protein interactions between enzyme and cofactor, structural changes upon complex formation, overview
-
putidaredoxin
cofactor putidaredxin-bound enzyme structure analysis using crystal structure, PDB ID 4JWS
-
putidaredoxin
Pdx, the enzyme is highly specific for its reductase, with an absolute requirement for the native putidaredoxin (Pdx) to provide the second electron transfer, binding kinetics and thermodynamics with wild-type and mutant enzymes, overview. Upon binding ferric P450cam, Pdx is now known to trigger a conformational change in the enzyme, which may provide the trigger to coordinate enzyme turnover and protect the enzyme from oxidative damage
-
putidaredoxin
the catalytic cycle of P450cam requires two electrons, both of which are donated by putidaredoxin (Pdx), a ferredoxin containing a [2Fe-2S] cluster, structures of the Pdx-P450cam complex, potential electron transfer pathways and interactions between Pdx Asp38 and P450cam Arg112, as well as hydrophobic contacts between the Pdx Trp106 and P450cam residues, favorable interactions exist between Pdx Tyr33 and P450cam Asp125, as well as between Pdx Ser42 and P450cam His352, overview
-
putidaredoxin
[2Fe-2S] putidaredoxin, Pdx. Camphor-bound ferric P450cam domain forms a complex with Pdx, modeling
-
cytochrome m
-
-
-
cytochrome m
-
cytochrome P-450cam, essential, b-type heme-thiolate protein of P-450-class
-
cytochrome m
-
optimal ratio of putidaredoxin reductase, putidaredoxin, cytochrome m is 1:10:2
-
cytochrome P450
-
cytochrome P450cam, importance and role of the polar residues, e.g. Tyr33, involved in the Pdx-P450cam interaction, overview. Interaction key residues are Pdx Asp38, Arg66, and Trp106, as well as P450cam Arg109 and Arg112, crystal structure of the Pdx-P450cam complex
-
NADH
-
-
NADH
-
requirement, reduces reductase flavoprotein and putidaredoxin, but not P450cam, in the absence of camphor
putidaredoxin
-
-
347695, 347697, 347698, 347699, 347700, 671535, 672760, 673037, 673071, 674152, 674438, 674474, 674959, 675234, 676280, 702301, 702820, 704713, 705331, 724338, 725602, 725873
-
putidaredoxin
-
essential, iron-sulfur redox protein, Fe2S2*Cys4-class, e-transfer agent to and effector of cytochrome P450cam
-
putidaredoxin
-
cannot be replaced by other FeS-proteins or the phospholipid of the hepatic microsomal P450 system
-
putidaredoxin
-
the putidaredoxin-binding site is only minimally affected by cytochrome b5
-
putidaredoxin
-
binding causes conformational changes involving K+, in absence of K+ multiple conformations occur, overview
-
putidaredoxin
-
electron transfer from putidaredoxin to the ferric-superoxo enzyme form, overview
-
putidaredoxin
-
the reaction cycle requires two distinct electron transfer processes from the [2Fe-2S] containing putidaredoxin to P450cam, altered binding and electron transfer with the putidaredoxin mutant C73S, structure and model of oxidized and reduced forms, overview
-
putidaredoxin
-
the substrate-free oxyferrous enzyme also reacts readily with reduced putidaredoxin
-
putidaredoxin
-
wild-type and mutant C73S/C85S cofactor, the mutant shows about 2fold improved substrate conversion
-
putidaredoxin
-
binding of putidaredoxin converts a single X-proline amide bond in CYP101 from trans or distorted trans to cis
-
putidaredoxin
-
optimal ratio of putidaredoxin reductase, putidaredoxin, cytochrome m is 1:10:2
-
putidaredoxin
-
transferring electrons from NADH to P450cam in a coupled assay method
-
putidaredoxin
-
the physiological electron transfer partner protein contains a [2Fe-2S] cluster, energetic importance and role of the polar residues, e.g. Tyr33, involved in the Pdx-P450cam interaction, overview. Interaction key residues are Pdx Asp38, Arg66, and Trp106, as well as P450cam Arg109 and Arg112, crystal structure of the Pdx-P450cam complex
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Fe2+
Iron
Fe2+
Fe3+
-
the enzyme requires K+ to drive formation of the characteristic high-spin state of the heme Fe3+ upon substrate binding
Iron
K+
Tl+
-
can substitute for K+ and minimize the effects of K+ absence on conformational perturbences upon putdaredoxin binding
Fe2+
heme iron, located in the active site, Thr101 is important for stability, tertiary and secondary structure of the heme active site, overview
Fe2+
Asn252 can stabilize the ferric hydroperoxy intermediate, preventing premature release of H2O2 and enabling addition of the second proton to the distal oxygen to generate the catalytic ferryl species
Fe2+
in putidaredoxin (Pdx), a ferredoxin containing a [2Fe-2S] cluster
Iron
purified reconstituted P450cam exhibits the ferrous CO-bound P450cam spectrum with the characteristic Soret band at 446 nm, indicating that the thiolate of Cys357 is ligated to the heme iron of the one-legged heme as seen in the wild-type protein
Fe2+
-
requirement, enzyme complex with two FeS-protein components
Fe2+
-
heme iron, formation of an oxyferrous cytochrome P450cam during reaction, structure of the active site of oxyferrous-P450cam
Fe2+
-
heme iron, heme centre tertiary structure and redox potential analysis of wild-type and C357M mutant enzymes, overview
Fe2+
-
the reaction cycle requires two distinct electron transfer processes from the [2Fe-2S] containing putidaredoxin to P450cam
Fe2+
-
local protein backbone dynamics of CYP101 depend upon the oxidation and ligation state of the heme iron
Iron
-
the hydroperoxo iron(III) intermediate P450camFeIII-OOH, i.e. Compound 0 involved in the natural catalytic cycle of P450cam, can be transiently observed in the peroxo-shunt oxidation of the substrate-free enzyme
K+
-
increase of activity, selective effector of enhanced substrate affinity to cytochrome m
K+
-
binding of K+ dramatically increases the high spin content for wild-type and mutant complexes of enzyme and substrate or substrate analogues
K+
-
effects of potassium ion binding on camphor-bound oxidized, camphor-bound reduced, and on CO-bound reduced wild-type and L358P enzyme, detailed overview, the enzyme requires K+ to drive formation of the characteristic high-spin state of the heme Fe3+ upon substrate binding, K+ binding site, overview
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
cyanide
binding of cyanide results in significant conformational changes and two different rotamers for residue D251
Metyrapone
enzyme P450cam bound to metyrapone is constrained in the closed conformation
additional information
-
increasing viscosity of the assay mixture, e.g. by addition of glucose or sucrose, reduces kcat
-
additional information
-
emulsions formed by strongly agitating the mixture with a vortexer denatured protein, P450cam has a complete loss of activity when incubated under these conditions for more than 5 min prior to the initiation of the reaction
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
bis(2-ethylhexyl) sulfosuccinate
-
improves the initial activity of P450cam in two-phase emulsions with initial camphor concentrations of 5-15 mM. P450cam is activated in the surfactant-free emulsions, and addition of bis(2-ethylhexyl) sulfosuccinate sodium salt improves the activity even further, at least over the range of camphor concentrations for which initial rates are readily measurable in all media. The largest rate enhancement is 4.5fold. Nearly 50times more product is formed in the surfactant-stabilized emulsions than is achieved in aqueous buffer
cumene hydroperoxide
-
considerable amount of the putative Cpd II species accumulates, even at pH 7.4. By contrast, reactions with meta-chloroperbenzoic acid at pH 7.4 yield very little of the ca. 420 nm species, unless methanol is included
methanol
-
reaction with meta-chloroperbenzoic acid in the presence of methanol (3% or ca. 1 M after mixing) at pH 7.4 and 25°C results in the accumulation of a considerable fraction of the P450cam as the putative Cpd II species
additional information
-
involvement of X-proline isomerization in enzyme function
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0046
3-chloroindole
mutant G120A/Y179H/G248S/D297H, pH 7.4, temperature not specified in the publication
0.016
3-chloroindole
mutant T56A/N116H/D297N, pH 7.4, temperature not specified in the publication
0.026
3-chloroindole
mutant Y179H, pH 7.4, temperature not specified in the publication
0.041
3-chloroindole
mutant G60S/Y75H, pH 7.4, temperature not specified in the publication
0.09
3-chloroindole
mutant D97F/P122L/Q183L/L244Q, pH 7.4, temperature not specified in the publication
0.15
3-chloroindole
mutant G93C/K314R/L319M, pH 7.4, temperature not specified in the publication
0.44
3-chloroindole
mutant E156G/V247F/V253G/F256S, pH 7.4, temperature not specified in the publication
additional information
additional information
kinetics of wild-type and mutant enzymes
-
additional information
additional information
combined quantum mechanical/molecular mechanical study, molecular dynamics, overview
-
additional information
additional information
spectroscopic and stopped flow transient kinetic studies
-
additional information
additional information
stopped-flow kinetic analysis of the peracid oxidation of the wild-type enzyme and substrate-free ferric mutant enzymes overview
-
additional information
additional information
thermal unfolding kinetics, substrate binding kinetics
-
additional information
additional information
detailed comparative pre-steady-state kinetic and steady-state analysis of enzyme mutant SeP450cam C357U/R365L/E366Q and its the cysteine-containing wild-type, overview
-
additional information
additional information
enzyme-cofactor interaction kinetics of wild-type and mutant enzymes, steady-state and stopped-flow reaction kinetics, overview
-
additional information
additional information
interaction thermodynamics between Pdx and various states of P450cam constrained in different conformations, overview
-
additional information
additional information
thermodynamics of wild-type and mutant enzymes with different substrates, overview
-
additional information
additional information
-
Km-value of putidaredoxin, 0.0038 mM, steady-state kinetics of enzyme interaction with redox partners
-
additional information
additional information
-
development of a global data fitting kinetic model that describes the time-varying concentrations of substrate and products, Michaelis-Menten kinetics
-
additional information
additional information
-
EPR/ENDOR studies of wild-type and mutant T252A enzymes
-
additional information
additional information
-
equilibrium isotope effect on O2 binding, isotope effects and transient-state kinetics, electron transfer kinetics, steady-state kinetics, recombinant His6-tagged enzyme, overview
-
additional information
additional information
-
single turnover kinetics
-
additional information
additional information
-
stopped-flow kinetics at different pH and temperature
-
additional information
additional information
-
stopped-flow kinetics of the reaction between putidaredoxin with 1,3-dimethoxy-5-methyl-1,4-benzoquinone, kinetics of the first and the second electron transfer to P450cam of wild-type and mutant enzymes
-
additional information
additional information
-
steady-state turnover activities of the P450cam catalytic cycle, overview
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
34
cytochrome m
-
P-450cam, constituent part of the multi-component enzyme
-
additional information
additional information
enzymatic substrate turnover with cytochrome b5 as the effector
-
0.0014
3-chloroindole
mutant Y179H, pH 7.4, temperature not specified in the publication
0.0053
3-chloroindole
mutant G120A/Y179H/G248S/D297H, pH 7.4, temperature not specified in the publication
0.011
3-chloroindole
mutant D97F/P122L/Q183L/L244Q, pH 7.4, temperature not specified in the publication
0.027
3-chloroindole
mutant T56A/N116H/D297N, pH 7.4, temperature not specified in the publication
0.06
3-chloroindole
mutant G60S/Y75H, pH 7.4, temperature not specified in the publication
0.062
3-chloroindole
mutant G93C/K314R/L319M, pH 7.4, temperature not specified in the publication
0.186
3-chloroindole
mutant E156G/V247F/V253G/F256S, pH 7.4, temperature not specified in the publication
17.9
NADH
pH 7.4, 22°C, recombinant mutant H352A enzyme with mutant S42A putidaredoxin
26.3
NADH
pH 7.4, 22°C, recombinant mutant D125A enzyme with mutant Y33A putidaredoxin
34.1
NADH
pH 7.4, 22°C, recombinant mutant H361A enzyme with wild-type putidaredoxin
34.1
NADH
pH 7.4, 22°C, recombinant wild-type enzyme with mutant S42A putidaredoxin
34.9
NADH
pH 7.4, 22°C, recombinant wild-type enzyme with mutant Y33A putidaredoxin
36.3
NADH
pH 7.4, 22°C, recombinant mutant H352A enzyme with wild-type putidaredoxin
37.4
NADH
pH 7.4, 22°C, recombinant wild-type enzyme with mutant S44Aputidaredoxin
38.2
NADH
pH 7.4, 22°C, recombinant wild-type enzyme with wild-type putidaredoxin
40.3
NADH
pH 7.4, 22°C, recombinant mutant D125A enzyme with wild-type putidaredoxin
56.4
reduced putidaredoxin
putidaredoxin reductase-camphor monooxygenase fusion enzyme, pH 7.4, 30°C
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.05
3-chloroindole
mutant Y179H, pH 7.4, temperature not specified in the publication
0.12
3-chloroindole
mutant D97F/P122L/Q183L/L244Q, pH 7.4, temperature not specified in the publication
0.42
3-chloroindole
mutant G93C/K314R/L319M, pH 7.4, temperature not specified in the publication
0.43
3-chloroindole
mutant E156G/V247F/V253G/F256S, pH 7.4, temperature not specified in the publication
1.15
3-chloroindole
mutant G120A/Y179H/G248S/D297H, pH 7.4, temperature not specified in the publication
1.49
3-chloroindole
mutant G60S/Y75H, pH 7.4, temperature not specified in the publication
1.68
3-chloroindole
mutant T56A/N116H/D297N, pH 7.4, temperature not specified in the publication
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.4
7.4
-
assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
22
25
37
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
20 - 47
-
tertiary structure of enzyme undergoes conformational changes in this temperature range in the absence of substrate
20 - 60
-
tertiary structure of enzyme undergoes conformational changes in this temperature range in the presence of substrate
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
-
Pseudomonas putida strains can use (1R)-(+) camphor as sole carbon and energy source
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
-
-
-
activity sediments after 14 h centrifugation of crude extract at 140000 * g, not after 2 h at 100000 * g, Pseudomonas sp. C5
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
cytochrome P450cam (CYP101) from Pseudomonas putida is the model enzyme for the P450 superfamily
malfunction
metabolism
putidaredoxin binds to a state of P450cam, where the substrate entry channel is partially open, the active site residues are positioned to facilitate the formation of a water-mediated proton relay network akin to those observed for the open state. The formation of this conformation is driven by binding of an allosteric camphor molecule prior to the first electron transfer step. Upon binding, the R186-D251 salt bridge is destabilized and breaks, allowing partial opening of the substrate entry channel and forming a conformation favorable to putidaredoxin binding. Putidaredoxin stabilizes this conformation during both electron transfer steps and keeps the R186-D251 bridge broken so a water-mediated proton relay network can form. Following the second electron transfer step, putidaredoxin remains in complex with P450cam and helps to promote the opening of channel 2
physiological function
cytochrome P450cam carries out the conversion of camphor to 5-exohydroxycamphor under conditions in which the bacterium uses camphor as the primary carbon source for growth
evolution
-
the enzyme belongs to the superfamily of cytochrome P450 monooxygenases
physiological function
-
Pseudomonas putida is capable of detoxification of camphor and borneol, overview
additional information
malfunction
alanine substitutions of the residues involved in putidaredoxin-enzyme interactions do not influence the rates of electron transfer, interaction analysis of enzyme wild-type and mutant (D125A, H352A, and H361A) proteins with putidaredoxxin wild-type and mutant (Y33A, S42A, and S44A) proteins, kinetics, overview
malfunction
structural, electronic, and catalytic properties of cytochrome P450cam are subtly altered when the cysteine that coordinates to the heme iron is replaced with a selenocysteine, mapping of the effects of the sulfur-to-selenium substitution on the individual steps of the catalytic cycle. The more electron-donating selenolate ligand has only negligible effects on substrate, product, and oxygen binding, electron transfer, catalytic turnover, and coupling efficiency. Off-pathway reduction of oxygen to give superoxide is the only step significantly affected by the mutation. Incorporation of selenium accelerates this uncoupling reaction approximately 50fold compared to sulfur, but because the second electron transfer step is much faster, the impact on overall catalytic turnover is minimal. Quantum mechanical calculations, Lewis structure of our P450cam active site model and quantum mechanically optimized structures (with TPSS/SVP) of the oxygen-bound forms with S and Se in the triplet spin state, overview
additional information
CYP101A1 is a soluble monomeric heme-containing camphor monooxygenase
additional information
-
CYP101A1 is a soluble monomeric heme-containing camphor monooxygenase
additional information
cofactor putidaredxin-bound and substrate-bound wild-type and E195C/A199C mutant enzymes, camphor-free, cyanide-bound P450cam, and apoenzyme structure analysis, structure analysis using crystal structure, PDB IDs 4JWS and 3L63. Cyanide and camphor can bind simultaneously in the active site
additional information
double electron-electron resonance measurements of intermolecular distances in the Pdx/P450cam complex show that the geometry of the complex is nearly identical for the open and closed states of P450cam
additional information
structures of the Pdx-P450cam complex, potential electron transfer pathways and interactions between Pdx Asp38 and P450cam Arg112, as well as hydrophobic contacts between the Pdx Trp106 and P450cam residues, favorable interactions exist between Pdx Tyr33 and P450cam Asp125, as well as between Pdx Ser42 and P450cam His352, overview
additional information
-
NMR structure analysis of the enzyme CYP101A1 with substrate (+)-camphor bound the active site and substrate-free enzyme after removal of (+)-camphor, molecular dynamics simulations and modeling, overview. Portions of a beta-rich region adjacent to the active site shift so as to partially occupy the vacancy left by removal of substrate. The accessible volume of the active site is reduced in the substrate-free enzyme relative to the substrate-bound structure
additional information
-
P450cam from Pseudomonas putida oxidizes (1R)-(+)-camphor to 5-exo-hydroxy camphor and further to 5-oxo-camphor
additional information
-
substrate recognition and selectivity, enzyme-substrate interactions, NMR structrue analysis, Analysis of 1H,15N-TROSY-HSQC spectra of CYP-(+)-camphor-CO, CYP-adamantenone-CO and CYP-norcamphor-CO. overview. Replacing the native substrate camphor with adamantanone or norcamphor causes perturbations in NMR-detected NH correlations assigned to the network,which includes portions of a beta-sheet and an adjacent helix that is remote from the active site
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
-
putidaredoxin reductase: MW 43000 Da, putidaredoxin: MW 12500 Da, cytochrome P450cam: MW 45000 Da, Pseudomonas putida PpG786, ultracentrifugal, diffusion and amino acid analysis
additional information
-
putidaredoxin reductase: MW 45000 Da, putidaredoxin: MW 11000 Da, cytochrome m: MW 44000 Da, Pseudomonas putida PpG786, gel filtration
additional information
-
FMN-containing NADH-reductase, MW 43500 Da, putidaredoxin, MW 12500 Da, cytochrome P-450cam, MW 45000 Da, Pseudomonas putida C1, ultracentrifugal analysis, gel filtration, end group analysis and quantification of prosthetic group material
additional information
-
putidaredoxin reductase, MW 43500 Da, putidaredoxin, MW 11594 Da, cytochrome m, MW 45000 Da, Pseudomonas putida PpG786, analytical data
additional information
-
multi-component mixed function oxidase, consisting of cytochrome P-450, MW 40000 Da, putidaredoxin and putidaredoxin reductase, slightly smaller than cytochrome P-450, Pseudomonas C1, gel filtration
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
?
x * 47561.6, recombinant mutant R365L/E366Q, mass spectrometry, x * 47607.4, recombinant mutant C357U, mass spectrometry
additional information
additional information
release of the substrate is caused both due to increased solubility of the substrate in solution in presence of alcohol and due to change in the tertiary structure of the active site of the enzyme, addition of alcohols to cytochrome P450cam causes a small change in the secondary structural elements but a significant change in the tertiary structural organization of the enzyme
additional information
secondary structure and localization of perturbation sites of cytochrome b5 and putidaredoxin, overview
additional information
structural changes upon complex formation between enzyme and cofactor cause an increase in beta-sheets and alpha-helix content, a decrease in population of random coil/3 10-helix structure, a redistribution of turn structures within the interacting proteins and changes in the protonation states or hydrogen-bondonh of amino acid carboxylic side chains
additional information
-
structural changes upon complex formation between enzyme and cofactor cause an increase in beta-sheets and alpha-helix content, a decrease in population of random coil/3 10-helix structure, a redistribution of turn structures within the interacting proteins and changes in the protonation states or hydrogen-bondonh of amino acid carboxylic side chains
additional information
tertiary and secondary structure of the heme active site, overview
additional information
the camphor-free structure is in an open conformation characterized by a water-filled channel created by the retraction of the F and G helices, disorder of the B' helix, and loss of the K+ binding. site. Presence of K+ alone does not alter the open conformation, while camphor alone is sufficient for closure of the channel
additional information
-
the camphor-free structure is in an open conformation characterized by a water-filled channel created by the retraction of the F and G helices, disorder of the B' helix, and loss of the K+ binding. site. Presence of K+ alone does not alter the open conformation, while camphor alone is sufficient for closure of the channel
additional information
three-dimensional structure and dynamics of the open and closed states of CYP101A1, NMR analysis and molecular dynamics simulations, overview
additional information
-
three-dimensional structure and dynamics of the open and closed states of CYP101A1, NMR analysis and molecular dynamics simulations, overview
additional information
-
multi-component mixed function monooxygenase consisting of putidaredoxin reductase, MW 47000, putidaredoxin, MW 11700, cytochrome m, MW 50500, Pseudomonas putida PpG786, SDS-PAGE
additional information
-
cytochrome P450cam dimerizes via the formation of an intermolecular disulfide bond
additional information
-
secondary structure analysis of wild-type and mutant C357M enzymes, overview
additional information
-
wild-type dioxygen complex structure: high occupancy and a ordered structure of the iron-linked dioxygen and two 'catalytic' water molecules that form part of a proton relay system to the iron-linked dioxygen
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
glycoprotein
-
small amount of carbohydrate in putidaredoxin and in cytochrome P-450cam
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
atomistic simulations study. The diffusion of camphor along the pathway near the substrate recognition site is thermodynamically preferred. The diffusion near the substrate recognition site is triggered by a transition from a heterogeneous collection of closed ligand-bound conformers to the basin comprising the open conformations of cytochrome P450cam. The accompanying conformational change includesthe retraction of the F and G helices and the disorder of the B' helix
camphor-free or camphor-bound P450cam mutant C334A in the absence of substrate and at high and low K+ concentration, protein in 50 mM Tris, pH 7.4, with or without 2-4 mM camphor, mixed with crystallization solution containing 50 mM Tris, pH 7.4, and 12-22% PEG 8000 with and without 200 mM K+, sitting drop vapour diffusion method, 6°C, X-ray diffraction structure determination and analysis at 1.50-1.79 A resolution
docking simulations for binding of 3-chloroindole, the wild-type does not accommodate 3-chloroindole in the active site, whereas all the mutants do. The mutants did not differ significantly in the Fe-N, Fe-C-2 or Fe-C-3 distances for 3-chloroindole
double electron-electron resonance studies. The geometry of the complex is nearly identical for the open and closed states of P450cam. Putaredoxin makes a single distinct interaction with its binding site on the enzyme and triggers the conformational change through very subtle structural interactions
fusion of putidaredoxin reductase PdR to the carboxy-terminus of camphor monooxygenase CYP101A1 (P450cam) via a linker peptide and reconstitution of camphor hydroxylase activity with free putidaredoxin gives a functional system with comparable in vivo camphor oxidation activity as the native system. In vitro, the fused systems steady state NADH oxidation rate is 2fold faster than that of the native system. In contrast to the native system, NADH oxidation rates for the fusion enzyme show nonhyperbolic dependence on putidaredoxin concentration
mutant Y96F/F87W/V247L, binding of substrate (+)-alpha-pinene in two orientations related by rotation of the molecule
purified recombinant His-tagged wild-type enzyme and enzyme mutants C357U and R365L/E366Q in complex with camphor, X-ray diffraction structure determination and analysis at 1.55-1.83 A resolution, molecular replacement
quantum mechanics and molecular mechanics study on the hydrogen abstraction reaction during hydroxylation of camphor in the quartet state. An energy barrier of 21.3 kcal/mol and a standard free energy of activation of 16.8 kcal/mol are obtained
recombinant wild-type enzyme and mutant L244A/C334A in complex with imidazole or 1-methylimidazole, hanging drop vapour diffusion method, purified recombinant protein in 50 mM potassium phosphate, 250 mM KCl, 50 mM DTT, and 36-52% ammonium sulfate, mixing with an equal volume of 0.001 ml of mother liquor containing 25 mM imidazole or 1-methylimidazole, room temperature, 2 days, cryoprotectant is 50 mM KCl, 25% ammonium sulfate and 30% glycerol, X-ray diffraction structure determination and analysis at 1.5-2.15 A resolution
the reconstituted P450cam at 1.8 A resolution reveals that the asymmetric one-legged heme is incorporated into the heme pocket in the same plane and in essentially the same conformation as the heme of the wild-type. A unique array of water molecules extending from the Tyr96 residue to the outside of the protein are present in the crystal structure
two ferric P450cam structures partially complexed with (+)-camphor, by sitting-drop vapour-diffusion method, at 1.3 A (soaked crystals) or 1.35 A (unsoaked crystals) resolution. Belongs to space group P43212, unsoaked crystals have unit-cell parameters of a = b = 63.38, c = 247.30, whereas soaked crystals have unit-cell parameters of a = b = 63.61 and c = 250.39. Structure of the unsoaked P450cam shows an active site that is partially occupied by (+)-camphor and a water molecule liganded to the haem iron and rotamers of Thr101. (+)-Camphor-bound form is the major component and the water-bound form is the minor component. In the soaked P450cam, the population of the major component increases, while the minor component decreases. (+)-Camphor binding induces rotation of Thr101 to form a hydrogen bond that acts as a hydrogen donor to a peripheral haem propionate. This bonding contributes to redox-potential change
mutant F87W/Y96F/V247L in complex with 1,3,5-trichlorobenzene or (+)-alpha-pinene
-
purified recombinant wild-type and D251N and T252A mutant enzymes in complex with O2, usage of a high pressure oxygen cell, a single crystal first is transferred into cryobuffer containing 50 mM Tris-HCl, pH 7.4, 0.4-0.6 M KCl, 1 mM D-camphor, 30% polyethylene glycol 4000, and 20% glycerol followed by reduction with 10 mM sodium dithionite for 10 min under anaerobic condition, soaking in the oxygen-saturated cryobuffer at -5°C for 5 min, X-ray diffraction structure determination and analysis at 1.55-2.10 A resolution
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
C136S
site-directed mutagenesis, putidaredoxin binding compared to wild-type
C285S
site-directed mutagenesis, putidaredoxin binding compared to wild-type
C334A
C357U
C357U/R365L/E366Q
site-directed mutagenesis, structural, electronic, and catalytic properties of cytochrome P450cam are subtly altered when the cysteine that coordinates to the heme iron is replaced with a selenocysteine, mapping of the effects of the sulfur-to-selenium substitution on the individual steps of the catalytic cycle. The more electron-donating selenolate ligand has only negligible effects on substrate, product, and oxygen binding, electron transfer, catalytic turnover, and coupling efficiency. Off-pathway reduction of oxygen to give superoxide is the only step significantly affected by the mutation. Incorporation of selenium accelerates this uncoupling reaction approximately 50fold compared to sulfur, but because the second electron transfer step is much faster, the impact on overall catalytic turnover is minimal. Quantum mechanical calculations, overview. Steady-state kinetic analysis revealed that the selenocysteine substitution has essentially no effect on the specific catalytic activity or the binding interaction with the electron donor Pdx, as both kcat and KM,Pdx are very similar for wild-type and mutant enzymes
C58S
site-directed mutagenesis, putidaredoxin binding compared to wild-type
C85S
site-directed mutagenesis, putidaredoxin binding compared to wild-type
D97F/P122L/Q183L/L244Q
mutant isolated by Sequence Saturation Mutagenesis, converts 3-chloroindole to isatin
E14C/S29C/C85S/C73S
site-directed mutagenesis, putidaredoxin binding compared to wild-type
E156G/V247F/V253G/F256S
mutant isolated by Sequence Saturation Mutagenesis, shows the highest maximal velocity in the conversion of 3-chloroindole to isatin
E195C/A199C/C334A
site-directed mutagenesis, substrate and cofactor binding of the mutant compared to the wild-type, overview
F87W/Y96F/L244A
enhanced binding and oxidation of (+)-alpha-pinene, production of 86% (+)-cis-verbenol + 5% (+)-verbenone
F87W/Y96F/L244A/V247L
enhanced binding and oxidation of (+)-alpha-pinene
G120A/Y179H/G248S/D297H
mutant isolated by Sequence Saturation Mutagenesis, converts 3-chloroindole to isatin
G248E
low catalytic activity, incubation with camphor, putidaredoxin reductase, and NADH results in partial covalent binding of heme to protein, pronase digestion of heme-bound protein releases 5-hydroxyheme
G60S/Y75H
mutant isolated by Sequence Saturation Mutagenesis, shows highest Km/kcat values for the conversion of 3-chloroindole to isatin
G93C/K314R/L319M
mutant isolated by Sequence Saturation Mutagenesis, converts 3-chloroindole to isatin
L244A/C334A
site-directed mutagenesis, mutation C334A prevents adventitious dimerization to facilitate crystallization but has no further effect on structure or activity of the enzyme, the L244A mutation leads to a highly increased Km and reduced activity for imidazole, but not for for 1-methylimidazole, and altered binding of imidazole to the active site and the active site heme involving residue Val247, overview
L244F/V247L
site-directed mutagenesis, the mutant exhibits moderate to high R-selectivity toward ethylmethylbenzene substrates and shows a narrow width of the binding pocket
L244N/V247L
site-directed mutagenesis, the mutant displays the highest S-selectivity toward substrates 1-ethyl-2-methylbenzene and 1-ethyl-3-methylbenzene, and low R-selectivity toward 1-ethyl-4-methylbenzene and shows a narrow width of the binding pocket
M184V/T185F
site-directed mutagenesis, the mutation introduces changes above the heme plane, prefers S-orientation of 1-ethyl-4-methylbenzene in the binding pocket of mutant, enantioselectivities of 1-ethyl-2-methylbenzene and 1-ethyl-3-methylbenzene are similar to the wild-type enzyme
Q227C
Q272C
R365L/E366Q
S190C
S190D
does not show any significant change in the rate constants of the substrate association, has almost no effect on the activation energy of substrate binding to the enzyme
S48C
T101V
site-directed mutagenesis, the mutant shows decreased thermal stability of the heme active site and reaction intermediates in the reaction, equilibrium unfolding compared to the wild-type enzyme
T192E
rate constants of the substrate association is much lower compared to the wild-type, activation energy for the substrate association is significantly higher in the T192E mutant compared to the S190D mutant or the wild-type enzyme
T252A
T252N
has comparable turnover number but higher Km value relative to the wild-type enzyme, due to a decrease in the camphor binding affinity, non-productive H2O2 generation is negligible
T252N/V253T
has comparable turnover number but higher Km value relative to the wild-type enzyme, due to a decrease in the camphor binding affinity, non-productive H2O2 generation is negligible
T56A/N116H/D297N
mutant isolated by Sequence Saturation Mutagenesis, shows highest Km/kcat values for the conversion of 3-chloroindole to isatin
Y179H
mutant isolated by Sequence Saturation Mutagenesis, converts 3-chloroindole to isatin
Y75F
the mutant shows an altered active site structure influencing catalysis
Y96C/C334A
site-directed mutagenesis, substrate recognition and binding compared to the wild-type, conformational selection mechanism
Y96F
Y96F/C334A
site-directed mutagenesis, substrate recognition and binding compared to the wild-type, conformational selection mechanism
Y96F/L244A/V247L
enhanced binding and oxidation of (+)-alpha-pinene, production of 55% (+)-cis-verbenol + 32% (+)-verbenone
Y96F/Y75F
the mutant shows an altered active site structure influencing catalysis
Y96N/C334A
site-directed mutagenesis, substrate recognition and binding compared to the wild-type, conformational selection mechanism
C334A
C357M
-
site-directed mutagenesis, comparison of the mutant structure to the wild-type one
D251N
-
site-directed mutagenesis, the mutant shows altered conformation of the I helix groove and misses the catalytically important water molecules in the dioxygen complex leading to lower catalytic activity and slower proton transfer to the dioxygen ligand compared to the wild-type enzyme
D38A
-
site-directed mutagenesis, the mutant shows altered electron transfer activity with higher Kd values for ferric P450cam and about 20% of the first electron transferring ability compared to the wild-type enzyme, the mutant forms a complex with 1,3-dimethoxy-5-methyl-1,4-benzoquinone
D38N
-
site-directed mutagenesis, the mutant shows altered electron transfer activity with higher Kd values for ferric P450cam and about 20% of the first electron transferring ability compared to the wild-type enzyme
F87W/Y96F/V247L
-
enhanced activity for oxidation of 1,3,5-trichlorobenzene or (+)-alpha-pinene, compared to wild-type, analysis of active-site structure, crystallization
G326A
-
site-directed mutagenesis in order to decrease the flexibility of the polypeptide at that point, spin state fractions with different substrates and compared to the wild-type enzyme. The mutant shows 40% reduced activity compared to the wild-type enzyme
I396A
-
site-directed mutagenesis, the substrate specificity is altered compared to the wild-type enzyme
I396G
-
site-directed mutagenesis, the substrate specificity is altered compared to the wild-type enzyme
I396V
-
site-directed mutagenesis, the substrate specificity is altered compared to the wild-type enzyme
L358P
M395I
-
site-directed mutagenesis, the substrate specificity is altered compared to the wild-type enzyme
M96Y
-
site-directed mutagenesis, the substrate specificity is altered compared to the wild-type enzyme
N244L
-
site-directed mutagenesis, the substrate specificity is altered compared to the wild-type enzyme
P89I
-
yields a mixture of both bound camphor orientations, that seen in putidaredoxin-free and that seen in putidaredoxin-bound CYP101. A mutation in CYP101 that destabilizes the cis conformer of the Ile-88-Pro-89 amide bond results in weaker binding of putidaredoxin
R66A
-
site-directed mutagenesis, reduced mutant electron transfer activity and increased Kd values for ferric P450cam compared to the wild-type enzyme
R66E
-
site-directed mutagenesis, reduced mutant electron transfer activity and increased Kd values for ferric P450cam compared to the wild-type enzyme
T101M
-
ratio of (R)- to (S)-1-phenylethanol produced from ethylbenzene is 89:11
T101M/T185F/V247M
-
ratio of (R)- to (S)-1-phenylethanol produced from ethylbenzene is 87:13
T185F
-
ratio of (R)- to (S)-1-phenylethanol produced from ethylbenzene is 78:22
T185L
-
ratio of (R)- to (S)-1-phenylethanol produced from ethylbenzene is 80:20
T185V
-
ratio of (R)- to (S)-1-phenylethanol produced from ethylbenzene is 74:26
T252A
T297D
-
site-directed mutagenesis, the substrate specificity is altered compared to the wild-type enzyme
V247A
-
ratio of (R)- to (S)-1-phenylethanol produced from ethylbenzene is 87:13
V247M
-
ratio of (R)- to (S)-1-phenylethanol produced from ethylbenzene is 83:17
V295I
-
ratio of (R)- to (S)-1-phenylethanol produced from ethylbenzene is 76:24
V87F
-
site-directed mutagenesis, the substrate specificity is altered compared to the wild-type enzyme
W106A
-
site-directed mutagenesis, the mutant shows altered electron transfer activity with higher Kd values for ferric P450cam and about 20% of the first electron transferring ability compared to the wild-type enzyme
W106F
-
site-directed mutagenesis, reduced mutant electron transfer activity and increased Kd values for ferric P450cam compared to the wild-type enzyme
Y29F
-
the cis conformer is destabilized by the absence of the hydrogen bond between the carbonyl oxygen of Ile-88 and the Tyr-29 hydroxyl group
Y33A
-
site-directed mutagenesis, reduced mutant electron transfer activity and increased Kd values for ferric P450cam compared to the wild-type enzyme
Y33F
-
site-directed mutagenesis, reduced mutant electron transfer activity and increased Kd values for ferric P450cam compared to the wild-type enzyme
Y75F
-
reaction with meta-chloroperbenzoic acid at 25°C, pH 8.0, is similar to that with the Y96F variant, although slightly more Cpd I (and possibly some Cpd ES) is present
Y96A
-
site-directed mutagenesis, the mutant gains the ability to hydroxylate indole to 3-hydroxyindole
Y96C
-
site-directed mutagenesis, the mutant gains the ability to hydroxylate indole to 3-hydroxyindole
Y96F
Y96F/Y75F
-
mutants produce changes in hydrogen bonding patterns and increase hydrophobicity that affect the ratio of heterolytic to homolytic pathways in reactions with cumene hydroperoxide, resulting in a shift of this ratio from 84/16 for wild-type to 72/28 for the Y96F/Y75F double mutant
Y96G
-
site-directed mutagenesis, the mutant gains the ability to hydroxylate indole to 3-hydroxyindole
Y96M
-
site-directed mutagenesis, the mutant gains the ability to hydroxylate indole to 3-hydroxyindole
Y96Q
-
site-directed mutagenesis, the mutant gains the ability to hydroxylate indole to 3-hydroxyindole
Y96S
-
site-directed mutagenesis, the mutant gains the ability to hydroxylate indole to 3-hydroxyindole
Y96T
-
site-directed mutagenesis, the mutant gains the ability to hydroxylate indole to 3-hydroxyindole
additional information
C334A
spectroscopically and enzymatically identical to wild-type, but does not form dimers in solution
C334A
the mutation of P450cam increases the protein stability compared to the wild-type enzyme
C334A
-
the mutation of P450cam increases the protein stability compared to the wild-type enzyme
C334A
site-directed mutagenesis, the mutation reduces protein aggregation, but has no effect on catalytic activity
C357U
site-directed mutagenesis, selenocysteine increases the affinity for oxygen 3-4-fold and accelerates the formation of superoxide 50fold, but the net effect of the C357U mutation on substrate hydroxylation is minimal because the second electron transfer step is much faster than superoxide formation under normal turnover conditions. As a consequence, selenocysteine is an excellent surrogate for the proximal cysteine in P450cam, maintaining both high monooxygenase activity and coupling efficiency
C357U
site-directed mutagenesis, the engineered gene contains the requisite UGA codon for selenocysteine, the sulfur-to-selenium substitution subtly modulates the structural, electronic, and catalytic properties of the enzyme. Catalytic activity decreases only 2fold, whereas substrate oxidation becomes partially uncoupled from electron transfer. The structure of mutant C357U, including the active site, is very similar to that of wild-type enzyme and mutant R365L/E366Q. The specific activity of the selenoenzyme mutant C357U is approximately half that of the mutant R365L/E366Q, which is 2fold less active than the wild-type enzyme
Q227C
site-directed mutagenesis, putidaredoxin binding compared to wild-type
Q272C
site-directed mutagenesis, putidaredoxin binding compared to wild-type
R365L/E366Q
site-directed mutagenesis, the structure of mutant R365L/E366Q is very similar to that of wild-type enzyme and mutant C357U. The specific activity of the selenoenzyme mutant C357U is approximately half that of the mutant R365L/E366Q, which is 2fold less active than the wild-type enzyme
S190C
mutation used for double electron-electron resonance studies. Residues S48C and S190C are at opposite ends of the substrate access channel to provide a longer distance measurement
S190C
site-directed mutagenesis, putidaredoxin binding compared to wild-type
S48C
mutation used for double electron-electron resonance studies. Residues S48C and S190C are at opposite ends of the substrate access channel to provide a longer distance measurement
S48C
site-directed mutagenesis, putidaredoxin binding compared to wild-type
T252A
about 5% of wild-type activity, similar spectra for oxyferrous mutant and wild-type except for Soret band position blue shifts. Epoxidation substrate 5-methylenylcamphor has a anomalous binding mode for the mutant
T252A
non-productive H2O2 generation is dominant, does not oxidize camphor, substrate binding affinity is similar to that of the wild-type enzyme
T252A
mutant displays an additional coupling pathway responsible for the epoxidation of 5-methylenylcamphor. During the reaction, camphor cannot prevent H2O2 release and hence the T252A mutant does not oxidize camphor
Y96F
the mutant shows an altered active site structure influencing catalysis
Y96F
site-directed mutagenesis, population and dynamics of the conformational states are largely unaltered by the Y96F mutation compared to the wild-type enzyme
C334A
-
identical with the wild-type monomer in terms of optical spectra, camphor-binding and turnover
C334A
-
is identical spectroscopically and enzymatically to wild-type but does not dimerize in solution at high concentrations
C334A
-
mutant, that is spectroscopically and enzymatically identical to wild-type CYP101 enzyme, but does not form dimers in solution, and so is more suitable for solution NMR studies than wild-type enzyme
C334A
-
site-directed mutagenesis, the C334A mutant is spectroscopically and enzymatically identical to the wild type but does not form dimers in solution, and so is more suitable for NMR structure analysis than the wild type enzyme
L358P
-
stereo- and regioselectivity for d-camphor hydroxylation unchanged
L358P
-
in absence of putidaredoxin, mutant shows ring-current signals typical for wild-type enzyme in presence of putidaredoxin, heme-environment of mutant mimics that of the putidaredoxin-bound wild-type, mutant accepts nonphysiological electron donors dithionite and ascorbic acid
L358P
-
site-directed mutagenesis, spin-state equilibrium in the L358P mutant is more sensitive to K+ than the wild-type enzyme
T252A
-
site-directed mutagenesis, the mutant does not show altered conformation of the I helix groove and the catalytically important water molecules in the dioxygen complex
T252A
-
site-sirected mutagenesis, comparison of substrate binding properties to the wild-type enzyme, overview
T252A
-
the mutant can epoxidize olefins like 5-methylenyl-camphor, but is ineffective in camphor hydroxylation
Y96F
-
reaction with peracetic acid at pH 8.0, 25°C, is similar to that with meta-chloroperbenzoic acid, except that even with 2.4 mM peracetic acid, all steps are slower than those with 0.150 mM meta-chloroperbenzoic acid
additional information
removal of heme-7-propionate decreases the (+)-camphor affinity by approximately 3fold, but exerts less of an influence on the other steps of the catalytic cycle of the monooxygenation reaction catalyzed by P450cam, does not exert an influence on the structure and electronic properties of the heme
additional information
construction of a fusion enzyme composed of enzyme P450cam and putidaroxin reductase, i.e. P450cam-PdR, kinetic model based on two-site binding of putidaredoxin by P450cam-PdR and inactive dimer formation of the fusion protein. Fusion co-expression with putidaredoxin results in a functional system with in vivo camphor oxidation activity comparable to the wild-type system , but P450cam-PdR is a class I P450 fusion protein that exhibits significantly more favorable catalytic behavior than that of the wild-type system. Further oxidation of 5-exo-hydroxycamphor to 5-oxo-camphor by the fusion enzyme is 39% lower than for the native system
additional information
enantioselectivity of a library of active-site mutants of chimeric P450cam-RhFRed towards the benzylic hydroxylation of structurally related regioisomers of ethylmethylbenzene, computational molecular modeling, overview
additional information
interaction analysis of enzyme wild-type and mutant (D125A, H352A, and H361A) proteins with putidaredoxxin wild-type and mutant (Y33A, S42A, and S44A) proteins, kinetics, overview
additional information
-
a facile way to significantly enhance the catalytic efficiency of the P450cam system by the coupling of its native electron transfer system with enzymatic NADH regeneration catalyzed by glycerol dehydrogenase in Escherichia coli whole cell biocatalysts, production of (+)-exo-5-hydroxycamphor and 5-keto-camphor, overview
additional information
-
construction of a catalytically active recombinant Escherichia coli whole cell biocatalyst harboring a cytochrome P450cam monooxygenase system from Pseudomonas putida coupled with enzymatic cofactor putidaredoxin mutant C73S/C85S regeneration, the cofactor mutant is 2fold more effective than the wild-type putidaredoxin
additional information
-
construction of the deletion mutant DELTA106, the mutant shows reduced electron transfer activity and increased Kd values for ferric P450cam compared to the wild-type enzyme
additional information
-
establishment of a functional CYP101-system in a cell-like aqueous compartment build by a stable water-oil emulsion with the non-ioninc surfactant tetraethylene glycol dodecyl ether in micro-scale, the enzyme is not active in an organic-aqueous biphasic system, e.g. with etahnol or glycol, incorporating an NADH-regeneration system using recombinant His-tagged bacterial glycerol dehydrogenase, method optimization, overview
additional information
-
in contrast to the wild-type, the more hydrophobic sites of the Tyr-to-Phe variants, compared to that of wild-type, favor homolysis more strongly and lead to considerable fractions of the Cpd II like species in reactions with peracids, especially at higher pH
additional information
-
establishment of an in vitro screening system for P450cam selecting variants by the activity on NADH
additional information
-
generation of a G327 insertion mutant in order to determine whether the length of the helix played a role in the sensitivity of the K' helix to substrate, the insertion mutant fails to express
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
thermal unfolding data, thermal multistep unfolding modell, Thr101 is important for thermal stability, equilibrium unfolding of wild-type C334A and mutant enzymes
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
2-mercaptoethanol retards loss of FeS-chromophore from putidaredoxin during purification
-
freeze-thawing, less than 5% loss of activity of cytochrome P450cam in the presence of camphor
-
glycerol, minimizes loss of FMN during purification of putidaredoxin reductase
-
multiple freezing and thawing, at -20°C, cytochrome m accumulates an equally active, heme-containing component of higher molecular weight, DTT reconverts it to native cytochrome m at 25°C
-
putidaredoxin suffers degradation by repeated cycles of freezing and thawing
-
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hexane
-
P450cam is stable and retains 80% of its initial activity after 1 h in a hexane/water emulsion at agitation speeds of less than 250 rpm
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
under aerobic conditions, cytochrome P450cam decays at 25°C with t1/2 of 180 min to cytochrome P420, not rapidly in the presence of camphor
-
347691
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80°C, 50 mM potassium phosphate buffer, pH 7.4, 50 mM KCl, 1 mM (+)-camphor
-196°C, no loss of activity of putidaredoxin reductase after repeated freeze-thaw cycles
-
0°C, putidaredoxin slowly loses its prosthetic group, 2-mercaptoethanol retards apoprotein formation
-
4°C, monomeric cytochrome P450cam can be stored at low protein concentrations for several days without appreciable accumulation of the dimer
-
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain XL-1 Blue by nickel affinity chromatography and repeated anion exchange chromatography
recombinant P450cam C334A from Escherichia coli NCM533. Protein purification includes a protamine sulfate cut to precipitate nucleic acids, and an ammonium sulfate cut to isolate CYP101A1
recombinant P450cam mutant C334A from Escherichia coli strain BL21(DE3) by anion exchange chromatography and gel filtration
recombinant wild-type and mutant enzymes from Escherichia coli by anion exchange chromatography and gel filtration to homogeneity
recombinant wild-type and mutant enzymes from Escherichia coli by anion exchange chromatography and hydrophobic interaction chromatography
recombinant wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by a process including anion exchange chromatography and gel filtration
recombinant wild-type enzyme and chimeric enzyme fusion protein P450cam-PdR from Escherichia coli strain BL21 (DE3)
soluble proteins separated by ultracentrifugation and purified using Ni2+-nitrilotriacetate column and gel filtration
native C334A enzyme mutant from Pseudomonas putida strain ATCC 17453 by ammonium sulfate fractionation, anion exchange chromatography, and gel filtration
-
recombinant C334A enzyme mutant from Escherichia coli strain NCM533 by ammonium sulfate fractionation, dialysis, anion exchange chromatography, and gel filtration
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene camC, expression of mutant enzymes in Escherichia coli strain BL21(DE3)
gene camC, recombinant expression of wild-type enzyme and chimeric enzyme fusion protein P450cam-PdR in Escherichia coli strain BL21 (DE3)
open reading frames for P450cam mutants including a 6 x His carboxyl-terminal tag cloned into the pCW(Ori+) expression vector using the NdeI and XbaI restriction sites, expressed in Escherichia coli
overexpression of P450cam mutant C334A in Escherichia coli strain BL21(DE3)
pCHC1 plasmid, encoding the wild type cytochrome P450cam (C334A mutant of the native enzyme). PCR products transformed into Escherichia coli XL1-blue super-competent cells. Wild-type and mutants expressed in Escherichia coli BL21 (DE3) cells
recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain XL-1 Blue from expression plasmids, pSUABC, under control of the salicylate promoter
recombinant expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
recombinant expression of wild-type and mutant enzymes in Escherichia coli, the selenoenzyme mutant SeP450cam C357U/R365L/E366Q is expressed in Escherichia coli strain XL1 blue cotransformed with plasmid pSUABC, whereas mutant R365L/E366Q P450cam and wild-type P450cam are expressed in a Escherichia coli strain BL21 Gold, which enhances the yields of the latter proteins but not of SeP450cam
recombinant expression of wild-type enzyme, mutant P450cam[Tyr96Phe]-RhFRed, and other enzyme mutants in Escherichia coli strain BL21(DE3)
expression of enzyme and cofactor, separately, in Escherichia coli strain DH5alpha
-
expression of enzyme and mutant cofactor in Escherichia coli strain BL21(DE3)
-
expression of wild-type enzyme, encoded by gene camC, with gene camB, encoding the putidaredoxin, in Escherichia coli strains BL21(DE3), expression of mutant enzymes in Escherichia coli strain DH10B
-
functional co-expression with glycerol dehydrogenase in Escherichia coli strain BL21(DE3)
-
functional expression in Escherichia coli DH5alpha of tricistronic constructs consisting of P450cam encoded by the first cistron and the auxiliary proteins, putidaredoxin and putidaredoxin reductase by the second and the third
-
individual expression of enzyme and cofactor putidaredoxin in Escherichia coli
-
mutant C334A transformed into chemically competent Escherichia coli NCM533 cells
-
mutants overexpressed from Escherichia coli strain NCM533 harboring modified pDNC334A plasmids that encode the appropriate mutant of CYP101 under control of the lac promoter
-
Pseudomonas putida, cam operon has been isolated, cloned and expressed in Escherichia coli, review
-
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
synthesis
biotechnology
-
bioengineered Escherichia coli cells possess a heterologous self-sufficient P450 catalytic system that may have advantages in terms of low cost and high yield for the production of fine chemicals
synthesis
additional information
synthesis
selective enzymatic oxidation of (+)-alpha-pinene to verbenol, verbenone, or myrtenol by enzyme mutants
synthesis
fusion of putidaredoxin reductase PdR to the carboxy-terminus of camphor monooxygenase CYP101A1 (P450cam) via a linker peptide and reconstitution of camphor hydroxylase activity with free putidaredoxin enables the production of the almost fully heme-incorporated CYP-FdR fusion that is catalytically active in vivo and in vitro
synthesis
-
camphor hydroxylation in reverse micelles depends on coexistence of enzyme with putidaredoxin, putidaredoxin reductase, and NADH, but not on H2O2
additional information
P450cam is not an antifungal target, but is an excellent model in which to study the phenomenon of drug resistance
additional information
knowledge of the conformational landscape is central to understanding P450 activity, which has important practical ramifications for the design of therapeutics with optimized pharmacokinetics, and the manipulation of P450s, and possibly other enzymes, for biotechnological applications
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AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Hedegaard, J.; Gunsalus, I.C.
Mixed function oxidation. IV. An induced methylene hydroxylase in camphor oxidation
J. Biol. Chem.
240
4038-4043
1965
Pseudomonas sp., Pseudomonas putida, Pseudomonas putida C1 / ATCC 17453
Tyson, C.T.; Lipscomb, J.D.; Gunsalus, I.C.
The role of putidaredoxin and P450 cam in methylene hydroxylation
J. Biol. Chem.
247
5777-5784
1972
Pseudomonas putida
Gunsalus, I.C.; Wagner, G.C.
Bacterial P-450cam methylene monooxygenase components: cytochrome m, putidaredoxin, and putidaredoxin reductase
Methods Enzymol.
52
166-188
1978
Pseudomonas putida
Gould, P.V.; Gelb, M.H.; Sligar, S.G.
Interaction of 5-bromocamphor with cytochrome P-450 cam. Production of 5-ketocamphor from a mixed spin state hemoprotein
J. Biol. Chem.
256
6686-6691
1981
Pseudomonas putida
Gelb, M.H.; Mlknen, P.; Sligar, S.G.
Cytochrome P450cam catalyzed epoxidation of dehydrocamphor
Biochem. Biophys. Res. Commun.
104
853-858
1982
Pseudomonas putida
Smith Eble, K.; Dawson, J.H.
Novel reactivity of cytochrome P-450-CAM. Methyl hydroxylation of 5,5-difluorocamphor
J. Biol. Chem.
259
14389-14393
1984
Pseudomonas putida
Sligar, S.G.; Filipovic, D.; Stayton, P.S.
Mutagenesis of cytochromes P450cam and b5
Methods Enzymol.
206
31-49
1991
Pseudomonas putida
Gelb, M.H.; Heimbrook, D.C.; Mlknen, P.; Sligar, S.G.
Stereochemistry and deuterium isotope effects in camphor hydroxylation by the cytochrome P450cam monoxygenase system
Biochemistry
21
370-377
1982
Pseudomonas putida
Tsai, R.L.; Gunsalus, I.C.; Dus, K.
Composition and structure of camphor hydroxylase components and homology between putidaredoxin and adrenodoxin
Biochem. Biophys. Res. Commun.
45
1300-1306
1971
Pseudomonas putida, Pseudomonas putida C1 / ATCC 17453
Katagiri, M.; Ganguli, B.N.; Gunsalus, I.C.
Reduction of palmitoyl dihydroxyacetone phosphate by mitochondria
J. Biol. Chem.
243
3542-3555
1968
Pseudomonas putida, Pseudomonas putida C1 / ATCC 17453
Peterson, J.A.; Ishimura, Y.; Griffith, B.W.
Pseudomonas putida cytochrome P-450: characterization of an oxygenated form of the hemoprotein
Arch. Biochem. Biophys.
149
197-208
1972
Pseudomonas putida, Pseudomonas putida PpG1
Yoshioka, S.; Takahashi, S.; Ishimori, K.; Morishima, I.
Roles of the axial push effect in cytochrome P450cam studied with the site-directed mutagenesis at the heme proximal site
J. Inorg. Biochem.
81
141-151
2000
Pseudomonas putida
Loida, P.J.; Sligar, S.G.
Engineering cytochrome P-450cam to increase the stereospecificity and coupling of aliphatic hydroxylation
Protein Eng.
6
207-212
1993
Pseudomonas putida
Hishiki, T.; Shimada, H.; Nagano, S.; Egawa, T.; Kanamori, Y.; Makino, R.; Park, S.Y.; Adachi, S.I.; Shiro, Y.; Ishimura, Y.
X-ray crystal structure and catalytic properties of Thr252Ile mutant of cytochrome P450cam: roles of Thr252 and water in the active center
J. Biochem.
128
965-974
2000
Pseudomonas putida
Schlichting, I.; Berendzen, J.; Chu, K.; Stock, A.M.; Maves, S.A.; Benson, D.E.; Sweet, R.M.; Ringe, D.; Petsko, G.A.; Sligar, S.G.
The catalytic pathway of cytochrome P450cam at atomic resolution
Science
287
1615-1622
2000
Pseudomonas putida
Unno, M.; Christian, J.F.; Sjodin, T.; Benson, D.E.; Macdonald, I.D.G.; Sligar, S.G.; Champion, P.M.
Complex formation of cytochrome P450cam with putidaredoxin: evidence for protein-specific interactions involving the proximal thiolate ligand
J. Biol. Chem.
277
2547-2553
2002
Pseudomonas putida
Nickerson, D.P.; Wong, L.L.
The dimerization of Pseudomonas putida cytochrome P450cam: practical consequences and engineering of a monomeric enzyme
Protein Eng.
10
1357-1361
1997
Pseudomonas putida
French, K.J.; Strickler, M.D.; Rock, D.A.; Rock, D.A.; Bennett, G.A.; Wahlstrom, J.L.; Goldstein, B.M.; Jones, J.P.
Benign synthesis of 2-ethylhexanoic acid by cytochrome P450cam: enzymatic, crystallographic, and theoretical studies
Biochemistry
40
9532-9538
2001
Pseudomonas putida
Jones, J.P.; O'Hare, E.J.; Wong, L.L.
Oxidation of polychlorinated benzenes by genetically engineered CYP101 (cytochrome P450(cam))
Eur. J. Biochem.
268
1460-1467
2001
Pseudomonas putida
Harford-Cross, C.F.; Carmichael, A.B.; Allan, F.K.; England, P.A.; Rouch, D.A.; Wong, L.L.
Protein engineering of cytochrome p450(cam) (CYP101) for the oxidation of polycyclic aromatic hydrocarbons
Protein Eng.
13
121-128
2000
Pseudomonas putida
Yoshioka, S.; Takahashi, S.; Hori, H.; Ishimori, K.; Morishima, I.
Proximal cysteine residue is essential for the enzymatic activities of cytochrome P450cam
Eur. J. Biochem.
268
252-259
2001
Pseudomonas putida
Sibbesen, O.; Zhang, Z.; Ortiz de Montellano, P.R.
Cytochrome P450cam substrate specificity: relationship between structure and catalytic oxidation of alkylbenzenes
Arch. Biochem. Biophys.
353
285-296
1998
Pseudomonas putida
Lee, D.S; Park, S.Y.; Yamane, K.; Obayashi, E.; Hori, H.; Shiro, Y.
Structural characterization of n-butyl-isocyanide complexes of cytochromes p450nor and P450cam
Biochemistry
40
2669-2677
2001
Pseudomonas putida (P00183), Pseudomonas putida
Lo, K.K.; Wong, L.L.; Hill, A.O.
Surface-modified mutants of cytochrome p450cam enzymatic properties and electrochemistry
FEBS Lett.
451
342-346
1999
Pseudomonas putida
Sono, M.; Perera, R.; Jin, S.; Makris, T.M.; Sligar, S.G.; Bryson, T.A.; Dawson, J.H.
The influence of substrate on the spectral properties of oxyferrous wild-type and T252A cytochrome P450-CAM
Arch. Biochem. Biophys.
436
40-49
2005
Pseudomonas putida (P00183)
Bell, S.G.; Chen, X.; Xu, F.; Rao, Z.; Wong, L.L.
Engineering substrate recognition in catalysis by cytochrome P450cam
Biochem. Soc. Trans.
31
558-562
2003
Pseudomonas putida
Pochapsky, S.S.; Pochapsky, T.C.; Wei, J.W.
A model for effector activity in a highly specific biological electron transfer complex: the cytochrome P450(cam)-putidaredoxin couple
Biochemistry
42
5649-5656
2003
Pseudomonas putida
Purdy, M.M.; Koo, L.S.; Ortiz de Montellano, P.R.; Klinman, J.P.
Steady-state kinetic investigation of cytochrome P450cam: interaction with redox partners and reaction with molecular oxygen
Biochemistry
43
271-281
2004
Pseudomonas putida
Limburg, J.; LeBrun, L.A.; Ortiz de Montellano, P.R.
The P450cam G248E mutant covalently binds its prosthetic heme group
Biochemistry
44
4091-4099
2005
Pseudomonas putida (P00183)
Bell, S.G.; Chen, X.; Sowden, R.J.; Xu, F.; Williams, J.N.; Wong, L.L.; Rao, Z.
Molecular recognition in (+)-alpha-pinene oxidation by cytochrome P450cam
J. Am. Chem. Soc.
125
705-714
2003
Pseudomonas putida (P00183), Pseudomonas putida
Wei, J.Y.; Pochapsky, T.C.; Pochapsky, S.S.
Detection of a high-barrier conformational change in the active site of cytochrome P450cam upon binding of putidaredoxin
J. Am. Chem. Soc.
127
6974-6976
2005
Pseudomonas putida (P00183)
Tosha, T.; Yoshioka, S.; Ishimori, K.; Morishima, I.
L358P mutation on cytochrome P450cam simulates structural changes upon putidaredoxin binding: the structural changes trigger electron transfer to oxy-P450cam from electron donors
J. Biol. Chem.
279
42836-42843
2004
Pseudomonas putida
Murugan, R.; Mazumdar, S.
Role of substrate on the conformational stability of the heme active site of cytochrome P450cam: effect of temperature and low concentrations of denaturants
J. Biol. Inorg. Chem.
9
477-488
2004
Pseudomonas putida
Deprez, E.; Gill, E.; Helms, V.; Wade, R.C.; Hui Bon Hoa, G.
Specific and non-specific effects of potassium cations on substrate-protein interactions in cytochromes P450cam and P450lin
J. Inorg. Biochem.
91
597-606
2002
Pseudomonas putida
Ichinose, H.; Michizoe, J.; Maruyama, T.; Kamiya, N.; Goto, M.
Electron-transfer reactions and functionalization of cytochrome P450cam monooxygenase system in reverse micelles
Langmuir
20
5564-5568
2004
Pseudomonas putida
Mouri, T.; Michizoe, J.; Ichinose, H.; Kamiya, N.; Goto, M.
A recombinant Escherichia coli whole cell biocatalyst harboring a cytochrome P450cam monooxygenase system coupled with enzymatic cofactor regeneration
Appl. Microbiol. Biotechnol.
72
514-520
2006
Pseudomonas putida
Murugan, R.; Mazumdar, S.
Effect of alcohols on binding of camphor to cytochrome P450cam: spectroscopic and stopped flow transient kinetic studies
Arch. Biochem. Biophys.
455
154-162
2006
Pseudomonas putida (P00183)
Kuznetsov, V.Y.; Poulos, T.L.; Sevrioukova, I.F.
Putidaredoxin-to-cytochrome P450cam electron transfer: differences between the two reductive steps required for catalysis
Biochemistry
45
11934-11944
2006
Pseudomonas putida
Manna, S.K.; Mazumdar, S.
Role of threonine 101 on the stability of the heme active site of cytochrome P450cam: multiwavelength circular dichroism studies
Biochemistry
45
12715-12722
2006
Pseudomonas putida (P00183)
OuYang, B.; Pochapsky, S.S.; Pagani, G.M.; Pochapsky, T.C.
Specific effects of potassium ion binding on wild-type and L358P cytochrome P450cam
Biochemistry
45
14379-14388
2006
Pseudomonas putida
Purdy, M.M.; Koo, L.S.; Montellano, P.R.; Klinman, J.P.
Mechanism of O2 activation by cytochrome P450cam studied by isotope effects and transient state kinetics
Biochemistry
45
15793-15806
2006
Pseudomonas putida
Rui, L.; Pochapsky, S.S.; Pochapsky, T.C.
Comparison of the complexes formed by cytochrome P450cam with cytochrome b5 and putidaredoxin, two effectors of camphor hydroxylase activity
Biochemistry
45
3887-3897
2006
Pseudomonas putida (P00183)
Karyakin, A.; Motiejunas, D.; Wade, R.C.; Jung, C.
FTIR studies of the redox partner interaction in cytochrome P450: the Pdx-P 450cam couple
Biochim. Biophys. Acta
1770
420-431
2007
Pseudomonas putida (P00183), Pseudomonas putida
Mouri, T.; Kamiya, N.; Goto, M.
Increasing the catalytic performance of a whole cell biocatalyst harboring a cytochrome p450cam system by stabilization of an electron transfer component
Biotechnol. Lett.
28
1509-1513
2006
Pseudomonas putida
Koe, G.S.; Vilker, V.L.
Effect of oxygen on the dehalogenation of 1,2-dibromo-3-chloropropane by cytochrome P450cam (CYP101)
Biotechnol. Prog.
21
1119-1127
2005
Pseudomonas putida
Celik, A.; Speight, R.E.; Turner, N.J.
Identification of broad specificity P450CAM variants by primary screening against indole as substrate
Chem. Commun. (Camb. )
2005
3652-3654
2005
Pseudomonas putida
Swart, M.; Groenhof, A.R.; Ehlers, A.W.; Lammertsma, K.
Substrate binding in the active site of cytochrome P450cam
Chem. Phys. Lett.
403
35-41
2005
Pseudomonas putida (P00183)
-
Murugan, R.; Mazumdar, S.
Structure and redox properties of the haem centre in the C357M mutant of cytochrome P450cam
ChemBioChem
6
1204-1211
2005
Pseudomonas putida
Davydov, R.; Perera, R.; Jin, S.; Yang, T.C.; Bryson, T.A.; Sono, M.; Dawson, J.H.; Hoffman, B.M.
Substrate modulation of the properties and reactivity of the oxy-ferrous and hydroperoxo-ferric intermediates of cytochrome P450cam as shown by cryoreduction-EPR/ENDOR spectroscopy
J. Am. Chem. Soc.
127
1403-1413
2005
Pseudomonas putida
Altun, A.; Guallar, V.; Friesner, R.A.; Shaik, S.; Thiel, W.
The effect of heme environment on the hydrogen abstraction reaction of camphor in P450cam catalysis: a QM/MM study
J. Am. Chem. Soc.
128
3924-3925
2006
Pseudomonas putida (P00183)
Spolitak, T.; Dawson, J.H.; Ballou, D.P.
Reaction of ferric cytochrome P450cam with peracids: kinetic characterization of intermediates on the reaction pathway
J. Biol. Chem.
280
20300-20309
2005
Pseudomonas putida
Nagano, S.; Poulos, T.L.
Crystallographic study on the dioxygen complex of wild-type and mutant cytochrome P450cam. Implications for the dioxygen activation mechanism
J. Biol. Chem.
280
31659-31663
2005
Pseudomonas putida
Glascock, M.C.; Ballou, D.P.; Dawson, J.H.
Direct observation of a novel perturbed oxyferrous catalytic intermediate during reduced putidaredoxin-initiated turnover of cytochrome P-450-CAM: probing the effector role of putidaredoxin in catalysis
J. Biol. Chem.
280
42134-42141
2005
Pseudomonas putida
Michizoe, J.; Ichinose, H.; Kamiya, N.; Maruyama, T.; Goto, M.
Functionalization of the cytochrome P450cam monooxygenase system in the cell-like aqueous compartments of water-in-oil emulsions
J. Biosci. Bioeng.
99
12-17
2005
Pseudomonas putida
Spolitak, T.; Dawson, J.H.; Ballou, D.P.
Rapid kinetics investigations of peracid oxidation of ferric cytochrome P450cam: nature and possible function of compound ES
J. Inorg. Biochem.
100
2034-2044
2006
Pseudomonas putida (P00183)
Hirao, H.; Kumar, D.; Shaik, S.
On the identity and reactivity patterns of the "second oxidant" of the T252A mutant of cytochrome P450cam in the oxidation of 5-methylenenylcamphor
J. Inorg. Biochem.
100
2054-2068
2006
Pseudomonas putida
Zurek, J.; Foloppe, N.; Harvey, J.N.; Mulholland, A.J.
Mechanisms of reaction in cytochrome P450: hydroxylation of camphor in P450cam
Org. Biomol. Chem.
4
3931-3937
2006
Pseudomonas putida
Verras, A.; Alian, A.; de Montellano, P.R.
Cytochrome P450 active site plasticity: attenuation of imidazole binding in cytochrome P450(cam) by an L244A mutation
Protein Eng. Des. Sel.
19
491-496
2006
Pseudomonas putida (P00183)
Sakurai, K.; Shimada, H.; Hayashi, T.; Tsukihara, T.
Substrate binding induces structural changes in cytochrome P450cam
Acta Crystallogr. Sect. F
65
80-83
2009
Pseudomonas putida (P00183)
Kim, D.; Heo, Y.S.; Ortiz de Montellano, P.R.
Efficient catalytic turnover of cytochrome P450(cam) is supported by a T252N mutation
Arch. Biochem. Biophys.
474
150-156
2008
Pseudomonas putida (P00183)
Pochapsky, S.S.; Dang, M.; OuYang, B.; Simorellis, A.K.; Pochapsky, T.C.
Redox-dependent dynamics in cytochrome P450cam
Biochemistry
48
4254-4261
2009
Pseudomonas putida
Behera, R.K.; Mazumdar, S.
Roles of two surface residues near the access channel in the substrate recognition by cytochrome P450cam
Biophys. Chem.
135
1-6
2008
Pseudomonas putida (P00183)
Ryan, J.; Clark, D.
P450cam biocatalysis in surfactant-stabilized two-phase emulsions
Biotechnol. Bioeng.
99
1311-1319
2008
Pseudomonas putida
Kim, D.; Ortiz de Montellano, P.R.
Tricistronic overexpression of cytochrome P450cam, putidaredoxin, and putidaredoxin reductase provides a useful cell-based catalytic system
Biotechnol. Lett.
31
1427-1431
2009
Pseudomonas putida
Hayashi, T.; Harada, K.; Sakurai, K.; Shimada, H.; Hirota, S.
A role of the heme-7-propionate side chain in cytochrome P450cam as a gate for regulating the access of water molecules to the substrate-binding site
J. Am. Chem. Soc.
131
1398-1400
2009
Pseudomonas putida (P00183)
Spolitak, T.; Dawson, J.H.; Ballou, D.P.
Replacement of tyrosine residues by phenylalanine in cytochrome P450cam alters the formation of Cpd II-like species in reactions with artificial oxidants
J. Biol. Inorg. Chem.
13
599-611
2008
Pseudomonas putida
Altun, A.; Kumar, D.; Neese, F.; Thiel, W.
Multireference ab initio quantum mechanics/molecular mechanics study on intermediates in the catalytic cycle of cytochrome P450(cam)
J. Phys. Chem. A
112
12904-12910
2008
Pseudomonas putida
OuYang, B.; Pochapsky, S.S.; Dang, M.; Pochapsky, T.C.
A functional proline switch in cytochrome P450cam
Structure
16
916-923
2008
Pseudomonas putida
Lee, Y.T.; Wilson, R.F.; Rupniewski, I.; Goodin, D.B.
P450cam visits an open conformation in the absence of substrate
Biochemistry
49
3412-3419
2010
Pseudomonas putida (P00183), Pseudomonas putida
Asciutto, E.K.; Dang, M.; Pochapsky, S.S.; Madura, J.D.; Pochapsky, T.C.
Experimentally restrained molecular dynamics simulations for characterizing the open states of cytochrome P450cam
Biochemistry
50
1664-1671
2011
Pseudomonas putida (P00183), Pseudomonas putida
Hoffmann, G.; Boensch, K.; Greiner-Stoeffele, T.; Ballschmiter, M.
Changing the substrate specificity of P450cam towards diphenylmethane by semi-rational enzyme engineering
Protein Eng. Des. Sel.
24
439-446
2011
Pseudomonas putida, Pseudomonas putida PpG1
Asciutto, E.K.; Young, M.J.; Madura, J.; Pochapsky, S.S.; Pochapsky, T.C.
Solution structural ensembles of substrate-free cytochrome P450(cam)
Biochemistry
51
3383-3393
2012
Pseudomonas putida
Hiruma, Y.; Gupta, A.; Kloosterman, A.; Olijve, C.; Olmez, B.; Hass, M.A.; Ubbink, M.
Hot-spot residues in the cytochrome p450cam-putidaredoxin binding interface
ChemBioChem
15
80-86
2014
Pseudomonas putida
Prasad, B.; Rojubally, A.; Plettner, E.
Identification of camphor oxidation and reduction products in Pseudomonas putida: new activity of the cytochrome P450cam system
J. Chem. Ecol.
37
657-667
2011
Pseudomonas putida, Pseudomonas putida ATCC 17453
Dang, M.; Pochapsky, S.; Pochapsky, T.
Spring-loading the active site of cytochrome P450 cam
Metallomics
3
339-343
2011
Pseudomonas putida
Kelly, P.; Eichler, A.; Herter, S.; Kranz, D.; Turner, N.; Flitsch, S.
Active site diversification of P450cam with indole generates catalysts for benzylic oxidation reactions
Beilstein J. Org. Chem.
11
1713-1720
2015
Pseudomonas putida (P00183)
Vandemeulebroucke, A.; Aldag, C.; Stiebritz, M.; Reiher, M.; Hilvert, D.
Kinetic consequences of introducing a proximal selenocysteine ligand into cytochrome P450cam
Biochemistry
54
6692-6703
2015
Pseudomonas putida (P00183), Pseudomonas putida ATCC 12633 (P00183)
Basom, E.; Manifold, B.; Thielges, M.
Conformational heterogeneity and the affinity of substrate molecular recognition by cytochrome P450cam
Biochemistry
56
3248-3256
2017
Pseudomonas putida (P00183)
Liou, S.H.; Myers, W.K.; Oswald, J.D.; Britt, R.D.; Goodin, D.B.
Putidaredoxin binds to the same site on cytochrome P450cam in the open and closed conformation
Biochemistry
56
4371-4378
2017
Pseudomonas putida (P00183)
Kammoonah, S.; Prasad, B.; Balaraman, P.; Mundhada, H.; Schwaneberg, U.; Plettner, E.
Selecting of a cytochrome P450cam SeSaM library with 3-chloroindole and endosulfan - Identification of mutants that dehalogenate 3-chloroindole
Biochim. Biophys. Acta
1866
68-79
2018
Pseudomonas putida (P00183), Pseudomonas putida
Johnson, E.O.; Wong, L.L.
Partial fusion of a cytochrome P450 system by carboxy-terminal attachment of putidaredoxin reductase to P450cam (CYP101A1)
Catal. Sci. Technol.
6
7549-7560
2016
Pseudomonas putida (P00183)
Hiruma, Y.; Gupta, A.; Kloosterman, A.; Olijve, C.; Oelmez, B.; Hass, M.; Ubbink, M.
Hot-spot residues in the cytochrome P450cam-putidaredoxin binding interface
ChemBioChem
15
80-86
2014
Pseudomonas putida (P00183)
Eichler, A.; Gricman, .; Herter, S.; Kelly, P.; Turner, N.; Pleiss, J.; Flitsch, S.
Enantioselective benzylic hydroxylation catalysed by P450 monooxygenases characterisation of a P450cam mutant library and molecular modelling
ChemBioChem
17
426-432
2016
Pseudomonas putida (P00183)
Franke, A.; van Eldik, R.
Spectroscopic and kinetic evidence for the crucial role of compound 0 in the P450cam-catalyzed hydroxylation of camphor by hydrogen peroxide
Chemistry
21
15201-15210
2015
Pseudomonas putida
Wang, B.; Li, C.; Dubey, K.D.; Shaik, S.
Quantum mechanical/molecular mechanical calculated reactivity networks reveal how cytochrome P450cam and its T252A mutant select their oxidation pathways
J. Am. Chem. Soc.
137
7379-7390
2015
Pseudomonas putida (P00183)
Basom, E.; Spearman, J.; Thielges, M.
Conformational landscape and the selectivity of cytochrome P450cam
J. Phys. Chem. B
119
6620-6627
2015
Pseudomonas putida (P00183)
Lai, R.; Li, H.
Hydrogen abstraction of camphor catalyzed by cytochrome P450cam A QM/MM study
J. Phys. Chem. B
120
12312-12320
2016
Pseudomonas putida (P00183)
Aldag, C.; Gromov, I.A.; Garcia-Rubio, I.; von Koenig, K.; Schlichting, I.; Jaun, B.; Hilvert, D.
Probing the role of the proximal heme ligand in cytochrome P450cam by recombinant incorporation of selenocysteine
Proc. Natl. Acad. Sci. USA
106
5481-5486
2009
Pseudomonas putida (P00183), Pseudomonas putida ATCC 12633 (P00183)
Skinner, S.; Liu, W.; Hiruma, Y.; Timmer, M.; Blok, A.; Hass, M.; Ubbink, M.
Delicate conformational balance of the redox enzyme cytochrome P450cam
Proc. Natl. Acad. Sci. USA
112
9022-9027
2015
Pseudomonas putida (P00183)
Rydzewski, J.; Nowak, W.
Thermodynamics of camphor migration in cytochrome P450cam by atomistic simulations
Sci. Rep.
7
7736
2017
Pseudomonas putida (P00183)
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